Plants from Sea to Sky...

Plants from Sea to Sky Abstracts

Table of Contents Plenary Session 1 ................................................................................................................................. 2

CD Nelson Lecture ................................................................................................................................ 3 Plenary Session 2 ................................................................................................................................. 4

Education Forum .................................................................................................................................. 6

Plenary Session 3 ................................................................................................................................. 6 Carl Douglas Symposium ................................................................................................................... 7

Concurrent Oral Presentations ....................................................................................................... 9 Cell Biology I ...................................................................................................................................................... 9 Seeds to Sky .................................................................................................................................................... 12 Abiotic Stress I ............................................................................................................................................... 15 Nutrients and Metabolism ......................................................................................................................... 18 Sexual Reproduction ................................................................................................................................... 21 Specialized Metabolism I (Anatomy and Defence) ............................................................................ 24 Abiotic Stress II ............................................................................................................................................. 27 Biotic Stress .................................................................................................................................................... 31 Cell Biology II (Cell Walls) ......................................................................................................................... 34 Ecophysiology ................................................................................................................................................ 37 Technological Innovations ........................................................................................................................ 40 Specialized Metabolism II .......................................................................................................................... 43 Development .................................................................................................................................................. 46 Biochemistry .................................................................................................................................................. 49 Applied Plant Biology ................................................................................................................................. 52 Biotic Interactions ........................................................................................................................................ 55

Posters – Session 1 ............................................................................................................................. 59 Abiotic Stress Posters (101-‐AB to 120-‐AB) ......................................................................................... 59 Biotic Stress Posters (121-‐BS to 141-‐BS) ............................................................................................. 69 Cell Biology I Posters (142-‐CB to 151-‐CB) ........................................................................................... 80 Nutrients and Metabolism Posters (152-‐NM to 154-‐NM) ............................................................... 85 Sexual Reproduction Posters (155-‐SR to 160-‐SR) ............................................................................ 87

Posters – Session 2 ............................................................................................................................. 90 Applied Biology / Global Food Security Posters (199-‐AP to 210-‐AP) ......................................... 90 Biochemistry Posters (211-‐BC to 214-‐BC) ........................................................................................... 96 Biotechnology/Tech Innovations Posters (215-‐BT to 218-‐BT) .................................................... 98 Biotic Interactions Posters (219-‐BI to 227-‐BI) ................................................................................ 100 Technological Innovations Posters (226-‐BI to 227-‐BI) ................................................................. 103 Cell Wall Posters (228-‐CW to 235-‐CW) ............................................................................................... 105 Development Posters (236-‐DV to 248-‐DV) ........................................................................................ 109 Education Posters (249-‐ED to 250-‐ED) ............................................................................................... 116 Specialized Metabolism Posters (251-‐SM to 260-‐SM) ................................................................... 117

Plants from Sea to Sky Delegates ............................................................................................... 122

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Plenary Session 1 Wednesday July 5; 9:00 - 11:20 am; Forest Sciences Centre, Room 1005 Chair: Geoffrey Wasteneys, University of British Columbia

PLEN1.1: Mechanical and Chemical Signals in the Control of Arabidopsis Stem Cells

Elliot M. Meyerowitz

Howard Hughes Medical Institute and Division of Biology and Biological Engineering, California Institute of Technology

Experiments indicate that physical stress in the shoot apical meristem of Arabidopsis thaliana controls at least two aspects of cell biology – the cortical cytoskeleton, and the subcellular location of the PIN1 auxin efflux carrier. Cortical microtubules align in shoot apical meristem epidermal cells such that they are parallel to the principal direction of maximal stress when the stress is anisotropic. PIN1 is asymmetrically distributed in the plasma membranes of the same cells, with the highest amount in the membrane adjacent to the most stressed side wall. As cellulose synthase complexes use microtubules as tracks, one effect of cytoskeletal response is to reinforce walls in the maximal stress direction, thereby directing cell expansion to be orthogonal to the stress – creating a system that, through stress patterns, senses tissue shape, and through the effects of stress on cell wall synthesis, changes tissue shape. One effect of mechanically-determined PIN1 localization is redirection of auxin flow, which controls rates of cell expansion and position of new leaf and floral primordia in the meristem. Mechanical stress therefore creates a supracellular, tissue-wide feedback system that responds to and creates plant shape, and controls phyllotaxis through auxin flow control. This system has been amenable to computational modeling, leading to predictive models that show how the interplay of mechanical and chemical signals plays a central role in pattern formation and morphogenesis at the shoot apical meristem.

[email protected]

PLEN1.2: Why gels matter: roles for pectin and alginate in development of walled organisms

Siobhan A. Braybrook

University of California, Los Angeles Sainsbury Laboratory, University of Cambridge

In organisms with walled cells the cell wall acts as the final regulator of cell growth and shape change. Whether single celled or multi-cellular, the material changes in the cell wall required for growth must be controlled in space and time in order to produce a fit and functional organism. In plants, the cell wall consists of a cellulose re-enforced gel matrix comprised of pectin and hemi-cellulose. In brown algae, the cell wall has less cellulose and is instead a thicker matrix comprised of alginate and fucans. Currently, we are focusing of the role of cell wall gels 9pectin and alginate) in development of plants and algae. Our lab has been investigating the physical properties of cell walls, and their components, towards an understanding of how these properties might change to allow cell growth. These investigations involve the development of new in situ mechanical testing methods for cell walls (both natural and in material mimics), examining wall physical properties during growth and development, and investigating the underlying genetic and hormonal regulatory mechanisms specifying these changes. Our systems of study include plants and brown algae: the Arabidopsis thaliana hypocotyl and shoot apex, the Fucus serratus embryo, the Sargassum muticum apex, and the Nicotiana benthamiana leaf epidermis.

[email protected]

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PLEN1.3: The plant microbiome at the intersection of metabolism and defense

Cara Haney

Michael Smith Laboratory, The University of British Columbia, Canada Department of Microbiology & Immunology, The University of British Columbia, Canada Plant root-associated microbial communities (the “rhizosphere microbiome”) influence plant growth and defense. In order to provide these benefits, bacteria must successfully colonize their plants hosts. Colonization requires successful competition for plant nutrients while evading plant defenses. To identify bacterial colonization determinants in the Arabidopsis rhizosphere, we performed a super-saturating transposon mutagenesis (TnSeq) screen with the plant growth promoting bacterial strain Pseudomonas fluorescens WCS365. This screen identified 93 P. fluorescens genes with potential roles in rhizosphere fitness. We have confirmed and characterized a subset of these genes. This screen informs our understanding of the plant nutrients and defense responses encountered by bacteria in the plant rhizosphere.

Post colonization, root-associated bacteria can modulate agriculturally important traits like growth and disease resistance. Rhizosphere bacteria can induce systemic resistance (ISR) or susceptibility (ISS) against foliar pathogens. Pseudomonas strains that trigger ISS also promote Arabidopsis growth specifically under nitrogen-limiting conditions suggesting that the bacterial-induced growth promotion may in some instances come at the cost of defense. Using a comparative genomics approach between ISS and ISR strains, we have identified bacterial genes that correlate with the ISS phenotype. We are currently testing candidate bacterial genes for their role in modulating plant growth and defenses. Collectively, this work will inform our understanding of how the plant microbiome coordinates plant metabolic status and immunity.

[email protected]

CD Nelson Lecture Wednesday July 5:30 – 6:15 pm; Forest Sciences Centre, Room 1005 Chair: Anja Geitmann, McGill University

CDN1: Ubiquitin Proteasome System and Plant Response to Environmental Stress

Sophia Stone

Biology Department, Dalhousie University, Halifax NS Canada

The ubiquitin proteasome system (UPS) play a pervasive role in growth, development, and response to environmental stress in eukaryotes. The UPS attaches a chain of ubiquitin molecules to selected proteins, which are then delivered to the 26S proteasome for degradation. The covalent attachment of ubiquitin, a small highly conserved protein, to substrate proteins (referred to as ubiquitination) require the sequential action of three distinctive enzymes, E1, E2 and E3. A large family of E3s or ubiquitin ligases are the key components for governing substrate specificity. The most prevalent ubiquitin ligases in plants are the RING-type E3s, which are defined by the presence of a E2-binding RING domain. The Arabidopsis and rice genomes contain over 450 and 425 genes, respectively, that encode for RING domain containing proteins. Our research has focused on understanding the function of a subfamily of seven RING-type E3s that contain a series of ankyrin repeats (RING-ANK) used for substrate protein interaction. Despite similarity in domain architecture, our research shows that the RING-ANK E3s are involved in a diverse array of responses and developmental processes. Phenotypic screens of homozygous T-DNA insertion mutants and transgenic overexpressing plants have implicated these ubiquitin ligases in ethylene biosynthesis, abscisic acid and jasmonate signalling, pathogen defence, cell death, response to abiotic stresses including formate toxicity and iron deficiency. The isolation of endogenous substrates has contributed significantly to our understanding how the RING-ANK E3s are able to regulate such diverse processes. Identified substrates include transcription factors, protein kinases and metabolic enzymes. The identification of substrates has also allowed us to begin to investigate the underlying molecular mechanisms that modulate E3-substrate engagement.

[email protected]

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Plenary Session 2 Thursday July 6; 9:00 am – 11:20 am; Forest Sciences Centre, Room 1005

Chair: Jae-Hyeok Lee, University of British Columbia

PLEN2.1: Between a rock and a hard place: tales of trace metal homeostasis in Chlamydomonas

Sabeeha S Merchant1,2 1Department of Chemistry and Biochemistry, University of California, Los Angeles, USA and 2Institute for Genomics and Proteomics, University of California, Los Angeles, USA.

Trace elements like Cu, Mn, Fe and Zn are critical for life because they enable chemistries that are not readily possible with the functional groups found on proteins and nucleic acids. Their reactivity, which makes them useful in biology, also gives them the potential to be toxic intracellular if the chemistry is not controlled. Accordingly, there are mechanisms in every organism to regulate the uptake, distribution and storage of these elements. We have developed Chlamydomonas, a green alga in the green plant lineage, as a reference organism for understanding trace metal homeostasis, especially in the context of the biogenesis and maintenance of the bioenergetics membranes of respiration and photosynthesis. Each organism has an elemental quota based on the metabolic requirement for these elements in cellular constituents. When nutritional conditions preclude attainment of this quota, mechanisms for maintaining metabolism in face of specific deficiencies are brought into play. These elemental economy measures include replacement of individual metalloproteins to reduce the quota and re-distribution of the scarce metal from less essential to more essential proteins. The metal quota can also be disrupted genetically or by physiological manipulation of Chlamydomonas cells. For instance, during Zn limitation, Chlamydomonas hyperaccumulates Cu, dependent on the nutritional Cu sensor Crr1, but is functionally Cu-deficient. In this situation, the metal is in chemical excess but biologically invisible because it accumulates in an intracellular compartment. We have used fluorescence probes for Cu(I) as well as nano-secondary ion mass spectrometry to visualize these structures, which based on staining for pH and polyphosphate, appear to be lysosome-related compartments. X-ray absorption spectroscopy (XAS) is consistent with Cu(I) accumulation in an ordered structure. Cu isotope labeling demonstrates that cuprosome-sequestered Cu(I) becomes bioavailable with priority over extracellular Cu when homeostasis is restored, indicative of the cuprosome being a dynamic metal-storing structure in the cell. The structure also contains Ca and polyphosphate, and in situations of over-supply, Fe or Mn. Fe and Mn hyper-accumulation is blocked in vtc mutants that are affected in polyphosphate accumulation. The metal storing organelle is important for handling transitions between metal overload and deficiency, indicating its importance as a storage reservoir rather than an end point dumpster, raising questions about the mechanisms of selectivity of reservoir loading and unloading.

[email protected]

Chair: Patrick Martone, University of British Columbia

PLEN2.2: Top-down modification of bottom-up processes: nutrient recycling by consumers enhances algal growth in marine ecosystems

Matthew E. S. Bracken University of California, Irvine Understanding the roles and relative importance of top-down and bottom-up factors on primary producers is a central goal of ecology. At first glance, the effects of nutrients and herbivores on primary producers seem straightforward: nutrients enhance producer abundance, whereas herbivores consume producers, reducing abundance. However, herbivores’ effects on primary producers are not limited to consumption. For example, molluscan grazers on rocky shores enhance nutrient availability and algal recruitment, both of which have the potential to increase resource availability. Here, I describe two experiments in which I partitioned consumers’ top-down and bottom-up effects on marine algae. First, I show how community context mediates the top-down versus bottom-up effects of snails on rocky intertidal seaweeds. In particular, I highlight the necessity of considering

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simultaneous effects of grazers on multiple co-occurring seaweed species. Second, I show that positive effects of grazers on algae are not only associated with local-scale nutrient recycling. I manipulated grazer abundances at the level of whole tide pools but controlled access of those grazers to experimental plots within each pool. I found that grazers’ consumptive effects were paradoxically greatest in pools where grazer densities were reduced and that positive effects scaled with grazer abundance. Grazers benefitted algae not only by recycling nutrients but also by facilitating recruitment of algae into the system. Slime trails left by molluscs acted as adhesive traps for algae, enhancing settlement. Our understanding of the effects of consumers and nutrients on primary producers is incomplete without accounting for the top-down modification of bottom-up processes.

Matthew Bracken [email protected] Chair: Karen Tanino, University of Saskatchewan

PLEN2.3: Updates on health effects of berries - New modes of action linked to the modulation of the gut microbiota

Yves Desjardins, Ph.D., Agr. Institute of Nutrition and Functional Foods, Laval University, Québec City, Québec, Canada, GIV 0A6 It is now widely accepted that berries are providing many health benefits. Indeed, their regular consumption has been shown in animal pre-clinical and human clinical studies to prevent or reduce the incidence of many disorders and illnesses like cardiovascular diseases, obesity, metabolic syndrome, diabetes, cancer, and neurodegenerative diseases. Most berries are rich sources of vitamins and phytonutrients and they are especially rich in anthocyanins, phenolic acids, flavonoids, and hydro-soluble and condensed tannins (proanthocyanidins). These molecules display strong antioxidant activity in vitro, but probably not in vivo. To this effect, there is still much debate on the exact mode of action of these bioactive molecules on health. Once believed to prevent oxidative stress linked to diseases, it now appears that it is not the way these molecules act in the body. Some claim that strawberry phytochemicals may stimulate endogenous antioxidant enzymes activity and interact with molecular signalling involved in disease etiology. While this may be possible, the poor bioavailability and extensive phase I and II metabolisms of these polyphenols molecules preclude such an action. We have recently shown that berry polyphenols can strongly affect the gut microbiota and favour the development of a beneficial gut microbial community. In particular, they can stimulate the growth of a mucus inhabiting bacteria called Akkermansia municiphila, the anti-obesity bacteria. This bacteria has been shown to improve intestinal epithelium tightness and to reduce low-grade inflammation. We will thus present in this conference some results to explain how polyphenols may actually act as prebiotics, thereby improving health. [email protected]

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Education Forum Friday July 7; 8:45 – 9:30 am; FSC 1005

Chair: Emily Indriolo ED.1: Cannabis in the classroom – Education for a ‘growing’ industry Mathias Schuetz1,2 1Department of Biology, Kwantlen Polytechnic University; 2Department of Botany, University of British Columbia. Cannabis use for medical purposes has been legal in Canada for over 15 years, and cannabis use for recreational purposes is slated for legalization in July 2018. These changes in cannabis prohibition have already sparked what is projected to be a multibillion-dollar industry in Canada, and things are evolving quickly. Although a national public health and safety campaign has been touted as a priority that will accompany cannabis legalization, there has been no mention of education and training programs aimed at cannabis cultivation. Several high profile cannabis product recalls due to tainted cannabis (mold, bacteria, and banned fungicides/pesticides) have uncovered gaps in the understanding of how to grow cannabis with quality control regulations. This presents a tremendous opportunity for both research and education programs focusing on cannabis plant biology. These programs will benefit people working in the existing cannabis industry, future cannabis cultivation professionals, and also the general public that will have interests in home cultivation. This joint meeting of the Canadian Society of Plant Biologists and the Canadian Society for Horticultural Science provides an opportunity for plant scientists and educators to work together in building a framework for how cannabis plant education programs could be developed.

Plenary Session 3 Friday July 7; 9:30 -10:10 am; FSC, Room 1005 Chair: Geoffrey Wasteneys, University of British Columbia

PLEN3.1: A chemical genetic roadmap to improved tomato flavor Denise Tieman1, Marcio Resende1, Matias Kirst1, Sanwen Huang2, Harry Klee1. 1. University of Florida, Gainesville FL 32611 2. Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, No. 7, Pengfei Road, Dapeng District, Shenzhen 518124, China Modern commercial tomato varieties are substantially less flavorful than heirloom varieties. To understand and ultimately correct that deficiency, we quantified flavor-associated chemicals in 398 modern, heirloom and wild accessions. A subset of these accessions were evaluated in consumer panels, identifying the chemicals making the most important contributions to flavor and consumer liking. Modern commercial varieties contain significantly lower amounts of many of these important flavor chemicals than older varieties. Whole genome sequencing and a genome-wide association study allowed us to identify genetic loci affecting most of the target flavor chemicals, including sugars, acids and volatiles. Together, these results provide an understanding of the flavor deficiencies in modern commercial varieties and the information necessary for recovery of good flavor through molecular breeding. [email protected]

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Carl Douglas Symposium Friday July 7; 10:40am -12:15pm; FSC, Room 1005 Chair: Rob Guy, University of British Columbia

CDS.1: Embracing the complexity of nature – computational and genomic approaches for understanding the development and evolution of forest trees

Andrew Groover US Forest Service and University of California Davis

Forest biology is rich with fascinating examples of plant development and evolution, but forest trees have traditionally been difficult subjects for research at the molecular genetic level. In this presentation, I will give three examples of how next generation sequencing technologies can be paired with computational approaches to address complex developmental and evolutionary processes directly in forest trees. In the first example, we developed a model system for the study of wood formation based on the gravitropic response and “tension wood” development in woody stems of Populus. Using RNAseq data from wood forming tissues after gravitropic stimulation and other treatments, a co-expression approach was used to cluster genes into co-expression modules, which were functionally annotated and correlated with quantitative wood phenotypic traits, and further dissected to identify individual candidate regulatory genes. In a second example, we extended the co-expression approach to identify modules of co-expressed genes from woody tissues that are conserved across a broad sampling of angiosperm trees. These conserved modules represent putative ancestral mechanisms regulating wood formation in angiosperms, and are further described in terms of functional properties and associated biological processes. A final example is given by a new system for functional genomics in Populus using a large collection of irradiation hybrids carrying insertions and deletions of chromosomal segments. Experiments will be described in which complex traits can be genetically dissected using this germplasm, as well as identification of individual genotypes with morphological variation for wood formation.

[email protected]

CDS.2: Evolution of phenolic 3-hydroxylases in land plants: On the way to lignin biosynthesis. Annette V. Alber1,2, Alexandra Basilio Lopes3, Hugues Renault1, Martine Schmitt3, Danièle Werck-Reichhart1, Jürgen Ehlting2 1Institute of Plant Molecular Biology, CNRS, University of Strasbourg, France 2Department of Biology & Centre for Forest Biology, University of Victoria, BC, Canada 3Laboratoire d’Innovation Thérapeutique, UMR CNRS 7200, Illkirch, France Lignin is a defining feature of all vascular plants, but was its pathway invented only once during land plant evolution? Monolignols are synthesized through the phenylpropanoid pathway, alongside a plethora of soluble phenolics acting as bioactive compounds. The cytochrome P450 CYP98A family catalyzes the entry step into G- and S-monolignol biosynthesis using 4-coumaroyl-shikimate as a substrate in angiosperms. Other CYP98A family members hydroxylise distinct 4-coumaroyl-conjugates to bioactive compounds such as rosmarinic or chlorogenic acid.

Bryophytes contain a single CYP98A gene, but obviously do not use it for lignin biosynthesis. Lycopods also contain a single CYP98A, but utilize a distinct pathway towards monolignols. Based on extensive substrate utilization profiles of CYP98A members from across the plant lineage, we showed that CYP98As from bryophytes, lycopods, and also from ferns have a substrate profile that is clearly distinct from that of angiosperm CYP98As related to lignin biosynthesis. These non-seedplant CYP98As are essentially unable to utilize 4-coumaroyl-shikimate, but instead efficiently utilize other 4-coumaroyl-conjugates. This strongly indicates that lignin biosynthesis in ferns, as in lycopods, is distinct from the pathway known in angiosperms.

Conifer CYP98A display a broad range substrate utilization profile being able to utilize both 4-coumaroyl-shikimate as well as many other substrates. Only within the angiosperms numerous gene duplications and losses occurred. Independent duplications with the Salicaceae and the Amborellaceae independently gave rise to a shikimate-specific

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(i.e lignin-associated) isoform and a broad-range isoform in both Amborella trichopoda and Populus trichocarpa. Together, this suggests that coumaroyl-shikimate biosynthesis through CYP98As was recruited for lignin biosynthesis only in seed plants.

[email protected]

CDS.3: Protein Complex in Natural Rubber Biosynthesis in Lettuce (Lactuca sativa) Dae-Kyun RO,* Yang QU, Romit CHAKRABARTY, Moonhyuk KWON, Elysabeth REAVELL-ROY, Eun-Joo KWON Department of Biological Sciences, University of Calgary, Calgary, AB, Canada Natural rubber (NR) is a biopolymer made by condensations of thousands of isopentenyl diphosphate in cis-configurations on the priming molecule, farnesyl diphosphate. Almost all NRs are currently produced from the Brazilian rubber tree. Despite the importance of NR in chemical and medical industries, molecular mechanism of NR biosynthesis remains elusive partly due to the lack of model system to which molecular genetics can be applied. Lettuce (Lactuca sativa) is known to synthesize NR with >1 million Da average molecular weight. Hence we have established lettuce as a model system to study NR metabolism in plants. In our experiments using lettuce, contradictory to the traditional views and findings, 1) silencing rubber elongation factor homologs (proposed to be critical catalysts for NR biosynthesis) did not influence NR biosynthesis in lettuce, and 2) recombinant cis-prenyltransferase (CPT) did not show any catalytic activity and could not complement the yeast rer2 mutant that lacks yeast cis-prenyltransferase. Intriguingly, a proteomics analysis of lettuce latex identified a distantly related CPT homolog, named as CPT-like protein (CPTL). Silencing of CPTL in lettuce displayed a remarkable reduction of NR in latex, and cellular localizations and yeast-2-hybrid data suggested that CPTL interacts with CPT. However, CPT/CPTL protein complex could not synthesize high molecular weight NR polymer in in vitro assays. These results demonstrated that CPT/CPTL protein complex is necessary for NR biosynthesis, but unknown additional proteins or structural components are still required for proper NR biosynthesis in plants. [email protected]

CDS.4: Cracking a tough case: advances in our understanding of sporopollenin and the Arabidopsis pollen wall Teagen D. Quilichini1, A. Lacey Samuels2, Carl J. Douglas2 1. National Resource Council Canada, Saskatoon, SK 2. Department of Botany, University of British Columbia, Vancouver, BC The pollen wall is a complex, specialized cell wall that serves to protect the male gametophyte from terrestrial stress. The outer layer of the pollen wall is dominated by the robust sporopollenin biopolymer, which provides a durable and sculptured encasement around pollen grains. Sporopollenin and other components of the outer wall are contributed by sporophytic tapetal cells that are spatially separated from pollen grains as they develop inside the anther. Although the mechanisms that enable this recalcitrant biopolymer to form remain elusive, the application of molecular genetics to a number of sporopollenin biosynthesis and trafficking questions has facilitated pollen wall research progress over the past decade. This presentation will review contributions made by members of Carl Douglas’ laboratory to the field of pollen wall biology, with a focus on the export of sporopollenin from the tapetum in Arabidopsis thaliana. The presentation will conclude with our current model for sporopollenin synthesis, transport and assembly. [email protected]

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Concurrent Oral Presentations Forest Sciences Centre, 2424 Main Mall

Cell Biology I Wednesday July 5 1:30-3:00pm; FSC Room 1003 Chair: Sherryl Bisgrove, Simon Fraser University CB1.1: Endoplasmic Microtubule and CLASP Interaction in Arabidopsis Roots P. Yen Le, J. Christian Ambrose Department of Biology, The University of Saskatchewan, Saskatoon, SK, Canada We found that the Arabidopsis CLASP protein localizes to Endoplasmic Microtubule (EMT) attachment sites at the cell cortex in root tip division zone cells. EMTs were unable to attach to the cell cortex in clasp-1 mutants, and were thus unable to efficiently join into the cortical MT array. This EMT-cortex anchoring appears to stabilize the cytoplasm to prevent cytoplasmic streaming and premature vacuolation in these densely cytoplasmic cells. To our knowledge, this is the first report of a protein localized to EMT-cortex attachment sites, and our results reveal a new function for EMTs in promoting cytoplasmic stability in a vacuolated plant cells. [email protected]

CB1.2: How does the microtubule associated proteinEB1b modulate root responses to mechanical cues and gravity? Hae Ryoung KIM, Vita LAI, Sherryl BISGROVE Department of Biological Sciences, Simon Fraser University, B.C. When roots growing through the soil come in contact with impenetrable objects such as rocks and other debris they are able to grow around, rather than trying to force their way through. This obstacle avoidance behavior involves detecting and responding to both mechanical cues and gravity. The microtubule associated protein END BINDING 1b (EB1b) plays a role in this process; it belongs to a family of proteins known as +TIPs because of their localization to the growing ends of microtubules. In addition to microtubule binding, EB1b also interacts with an array of additional proteins that are involved in a wide array of processes in the cell. To further understand how EB1b modulates root responses to mechanical cues and gravity, we are investigating the biological importance of its two major activities: microtubule binding and interactions with other proteins. We have generated transgenic eb1b mutant lines expressing truncated versions of EB1b that are missing either microtubule-binding or the protein-protein interaction domains. Here, we provide evidence that the interaction of EB1b with other non-tubulin proteins is required while microtubule-binding of EB1b may not be needed, for normal root responses to mechanical cues and gravity. Hae Ryoung (Tommy) Kim / [email protected]

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CB1.3: Identification and characterization of a novel lipid droplet protein in Arabidopsis Michal PYC1*, Satinder K. GIDDA1, Yingqi CAI2, Olga YURCHENKO3, Kent D. CHAPMAN2, John M. DYER3, & Robert T. MULLEN1 1. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, N1G 2W1, Canada 2. Department of Biological Sciences, Center for Plant Lipid Research, University of North Texas, Denton, TX, 76203, USA 3. U.S. Department of Agriculture, Agricultural Research Service, U.S. Arid-Land Agricultural Research Center, Maricopa, AZ, 85138, USA Lipid droplets (LDs) are evolutionarily conserved organelles found in a wide range of unicellular and multicellular organisms. Uniquely enclosed by a single phospholipid monolayer and coated with a diverse set of proteins, LDs function primarily in the storage of energy-rich neutral lipids, such as triacylglycerols. In plants, LDs have been studied mostly in oilseeds, although all plant cell types, including those in vegetative tissues, have the machinery required for accumulating and storing lipids in LDs. We recently identified a class of proteins in Arabidopsis called the LD-associated proteins (LDAPs) that are abundant components of LDs in non-seed cell types and are required for their proper compartmentation. In an effort to identify other novel LD proteins, the LDAPs were used as ‘bait’ proteins in yeast two-hybrid screens with an Arabidopsis cDNA ‘prey’ library. Here, we describe one LDAP interactor, termed Arabidopsis mycobacterial-like protein (AMLP), which is homologous to members of a M. tuberculosis protein family involved in lipid transport. We show that, similair to the LDAPs, AMLP is expressed in Arabidopsis in a variety of tissues and at various developmental stages, and is targeted to the LD surface with high specificity. Furthermore, analyses of an Arabidopsis amlp mutant revealed (i) conspicuously enlarged LDs in both leaves and mature seeds, suggesting that AMLP plays a role in regulating LD size in plant cells, and (ii) altered neutral lipid levels in both leaves and seeds. Potential mechanisms by which AMLP functions in LD-related processes in plants will be discussed. Michal Pyc [email protected]

CB1.4: Mapping Polysaccharide Synthesis in a Changeable Golgi Apparatus

Miranda J. Meents1,2, Shawn D. Mansfield2, A. Lacey Samuels1 1 Department of Botany, University of British Columbia, Vancouver, Canada 2 Department of Wood Science, Faculty of Forestry, University of British Columbia, Vancouver, Canada The Golgi apparatus is central to endomembrane trafficking and essential for innumerable cellular processes. Despite this importance, our understanding of how the Golgi carries out these functions is surprisingly lacking in many cell types and developmental processes. In plants, cell wall synthesis is a fundamental function of the Golgi, and yet we do not know how the critical resident enzymes and cell wall polysaccharides are organized during synthesis and trafficking. To address this question, we quantitatively characterized the Arabidopsis Golgi apparatus in cells undergoing abundant secondary cell wall (SCW) production during xylem development. The onset of SCW synthesis coincided with a dramatic increase in the size of secretory vesicles and Golgi diameter, as quantified using transmission electron microscopy (TEM). Confocal microscopy of Golgi labelled with fluorescently-tagged resident proteins also showed a significant increase in Golgi density during this time. Next, we investigated the distribution of Golgi resident proteins and polysaccharide cargo by mapping their location in the Golgi stack using immuno-TEM. Both the SCW hemicellulose, xylan, and one of its biosynthetic enzymes, IRX9, were found in the intricate, fenestrated margins of the Golgi cisternae. This distribution of IRX9 was consistent with the ring-shaped Golgi signal of IRX9-GFP, as seen in confocal microscopy. Previous models of Golgi organization have hypothesized that Golgi residents reside in the compressed cisternal centers, and Golgi cargo in the swollen margins. Our data suggests otherwise, indicating the need for an amended model of Golgi organization.

Presenting Author: Miranda Meents ([email protected])

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CB1.5: Lunapark proteins suppress the membrane fusion activity of RHD3 for the formation of tubular ER network Jiaqi Sun*, Hugo Zheng Biology Department, McGill University, Montreal, QC, Canada The endoplasmic reticulum (ER) is a network of tubules and sheets playing important roles in eukaryotic cells. The formation of the ER requires homotypic membrane fusion, which is mediated by a Dynamin-like large GTPase protein family. In Arabidopsis, ROOT HAIR DEFECTIVE3 (RHD3) is an ER membrane fusogen, but little is known about how the activity of RHD3 is regulated in mediating the ER fusion. Here we show that Lunapark (LNP) proteins, a conserved ER membrane protein family, physically interact with RHD3 at 3-way junctions of the ER. Arabidopsis lunapark knockout mutants exhibit short root hairs with sheeted ER in many cells. We show that LNPs and RHD3 act antagonizingly in the fusion of ER membranes at the molecular and genetic levels. We thus suggest a model in which LNPs suppress the membrane fusion activity of RHD3 to regulate the formation of the ER network and cell growth. RHD3 mediates ER tubular membrane fusion to create 3-way junctions, after which LNPs are recruited to the 3-way junctions. LNPs recruited by RHD3 suppress the fusion activity of RHD3 so to stop further fusion thereby a 3-way junction can be stabilized. A loss of LNP activity would lead to an excessive ER fusion resulting a sheeted ER in cells. [email protected]

CB1.6: The emerging role of plant ER-PM contact sites as stress signaling platforms Abel Rosado Department of Botany, Faculty of Sciences. University of British Columbia, Vancouver, V6T 1Z4, Canada The coordination of segregated biochemical activities among eukaryotic organelles relies on evolutionarily conserved membrane micro-domains known as membrane contact sites (MCS). MCS are established and controlled by tethering structures that facilitate the transfer of molecules between closely apposed membranes. In plants, endoplasmic reticulum-plasma membrane contact sites (EPCS) are essential for the regulation of multiple stress and developmental responses. However, the molecular triggers of EPCS establishment and dynamics remain largely unknown. In this report, we describe an EPCS establishment mechanism where the cortical cytoskeleton (i) interacts with EPCS-localized Synaptotagmin (SYT) tethering complexes, (ii) is required for the delivery of SYT tethers to cortical ER subdomains, and (iii) regulates the establishment of SYT-labelled EPCS in response to environmental stresses. This mechanism highlights a plant-specific interplay between EPCS and cortical cytoskeleton, and provides a framework for the spatio-temporal regulation of plant stress responses. [email protected]

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Seeds to Sky Wednesday July 5 1:30-3:00pm; FSC Room 1001 Chair: Santokh Singh, University of British Columbia

SS.1: BSAS4;1- Key Candidate in Developing Common Bean as an Excellent Source of Protein Jaya JOSHI1, 2*, Justin B. RENAUD1, and Frédéric MARSOLAIS1, 2 1London Research and Development Centre, Agriculture and Agri-Food Canada, London, ON, N5V 4T3, Canada 2Department of Biology, University of Western Ontario, London, ON, N6A 3K7, Canada

With the increasing food insecurity in the populated world, the number of people affected by chronic undernourishment is also increasing. Protein energy malnutrition alone is causing 6 million deaths annually. Despite being a good source of protein and dietary fibre, quality of bean protein is questioned because of sub optimal levels of essential sulfur amino acids: methionine and cysteine. Levels of cysteine and methionine in developing seeds have an inverse relationship with the non-proteinogenic sulfur amino acid S-methylcysteine (SMC). One of the strategies to improve protein quality can be to redirect the sulfur from SMC to the cysteine pool. Elucidation of unknown biochemical pathway of SMC synthesis with 13C and 15N labelled serine and cysteine revealed serine as the precursor of SMC biosynthesis. Feeding common bean developing seeds with 13C and 15N lebelled methionine also suggested role of methionine in SMC biosynthesis. In cytoplasm, methanethiol released during hydrolysis of methionine might be condensed with O-acetylserine to form SMC. BSAS4;1, a cysteine synthase, utilizes O-acetylserine as a carbon backbone donor and methionine as a methyl donor to synthesize SMC. According to the Canadian Food Inspection Agency, the label “Good source of protein” can be used for beans and other legumes. In future silencing of SMC pathway candidate genes or development of TILLING lines with high cysteine and methionine levels may change bean from a good source to an “excellent source of protein”. Hence our findings would provide a helping hand to overcome the food insecurity in the growing world.

Jaya Joshi: [email protected]

SS.2: Making a better canola seed: Transcriptional and epigenetic profiling of Brassica napus seed development Deirdre KHAN1*, Michael G BECKER1, Isobel AP PARKIN2, Steven J ROBINSON2, Mark F BELMONTE1 1. Department of Biological Sciences, University of Manitoba, MB 2. Agriculture and Agrifoods Canada, SK Canola (Brassica napus) is one of Canada’s most economically valuable crops. The two most valuable products – canola oil and canola meal – are both derived from the seed. Improving canola seed quality through traditional breeding methods is both time-consuming and laborious, and we know very little about the genetic mechanisms that control valuable seed traits such as oil and protein content. To understand the transcriptional networks that underlie canola seed development, we used next generation RNA sequencing to study whole canola seeds at the ovule, globular, heart, mature green, and dry seed stages of seed development. Whole genome bisulfite sequencing was then used to profile the canola seed methylome at an early (globular) and late stage (mature green) of seed development. Early and late seed development form two distinct transcriptional clades. The methylomes of the globular and mature green stage seed are significantly distinct, with greater divergence in methylation at the ChG and CHH contexts than observed in the CpG context. Further, several bZIP transcription factors were identified as potential regulators of energy metabolism in early seed development, and bHLH transcription factors as putative regulators of embryo maturation and seed dormancy. Finally, we are using RNAi and overexpression to characterize key regulators in seed development, and the role they play in modulating seed protein and oil content. By understanding the patterns of DNA methylation and gene expression, we are finding new ways to probe seed development and improve the quality of canola seeds. Deirdre Khan [email protected]

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SS.3: Investigation of a novel seed-size regulating gene in Canola (Brassica napus) Trinh NGUYEN 1*, Soomin LEE 2 Chengbo YANG 3, Limin WU 4, Guanqun (Gavin) CHEN 1,5 1. Department of Biological Sciences, University of Manitoba, Winnipeg, Manitoba, Canada 2. St. John's Ravenscourt School, Winnipeg, Manitoba, Canada 3. Department of Animal Science, University of Manitoba, Winnipeg, Manitoba, Canada 4. InnoTech Alberta, Vegreville, Alberta, Canada 5. Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta, Canada Canola generates $26.7 billion in revenue annually in Canada; it is used in different food production and industrial processes. It is estimated that a 1% absolute increase in seed oil content would translate into an annual increase of $90 million. Various efforts have been made to increase oil yield to meet the world’s vegetable oil demand. One potential approach is to properly increase seed size in order to have more space for oil accumulation while keeping the seed size suitable for the current commercial equipment for canola seed processing. We recently generated a canola mutant with 28% smaller seeds by random T-DNA insertions. The preliminary segregation analysis showed that the smaller phenotype might be caused by over-expression of one unknown gene. Therefore, we hypothesize that a knockout mutation of this unknown gene would result in bigger seeds. The objective of this study is to identify and validate this gene and its function. Through our recent experiments, the partial sequence of the target gene was found via genome walking and six candidate genes were identified in the canola genome database. The subsequent gene expression analysis indicated that one gene had a higher expression in mutant flowers, pods and seeds when compared to the wild type (P < 0.05). The function of the homolog gene in Arabidopsis thaliana seed development is under evaluation. The results from this experiment will expand our understanding of seed development and provide a novel gene target allowing genetic modification via CRISPR/Cas9 to improve canola oil production. Trinh Thi Viet Nguyen [email protected]

SS.4: Histology and biochemistry of resin vesicles in conifer seeds Kristina KSHATRIYA1, 2*, Justin G.A. WHITEHILL1, Jörg BOHLMANN1, 2 1. Michael Smith Laboratories, University of British Columbia, BC 2. Department of Forest and Conservation Sciences, University of British Columbia, BC Canadian reforestation programs rely on orchards for seed production of most conifer species. The seeds of several economically important conifers contain an abundance of terpene rich resin vesicles that reduce germination of a seedlot when ruptured. Resin vesicles form on the outer layers of seeds and contain oleoresin. These structures are easily damaged during handling and processing for storage preparation, which significantly reduces the germination of a seedlot. The ecological and biological role of these specialized terpene-containing structures is largely unknown. We hypothesize that resin vesicles function in defense against seed feeding pests in addition to a potential role in germination biology. We evaluated the histological and biochemical features of resin vesicles from resin vesicle containing conifer species found throughout Canada. Our lab has developed a unique set of genomic resources for western redcedar. Therefore, we are focusing on resin vesicle development in western redcedar to identify resin vesicle biogenesis genes. Resin vesicles develop between the sarcotesta and the sclerotesta - two layers of the seed coat. Epithelial cells line the inside of resin vesicles, similar to axial resin ducts in cortex of young conifer stems. GC-MS analysis of resin vesicle containing seeds revealed a diversity of terpenes. Compound abundance and quality varied largely with seedlot and species. Western redcedar cones were monitored during the growing season to identify crucial timepoints in resin vesicle biogenesis. The ultimate goal of this project is to improve seed handling methods and selection of higher germinating seedlots based on terpene profiling approaches. Kristina Kshatriya - [email protected]\

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SS.5: Plant DGAT1 variants with enhanced performance generated by directed evolution Yang XU1*, Guanqun CHEN1, Michael S. GREER1, Kristian Mark. P. CALDO1,2, Geetha RAMAKRISHNAN1, Saleh SHAH1, Limin WU3, M. Joanne LEMIEUX2, Jocelyn OZGA1, and Randall J. WESELAKE1 1. Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta, Canada 2. Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada 3. Crop Pathology and Biotechnology, InnoTech Alberta, Vegreville, Alberta, Canada Diacylglycerol acyltransferase (DGAT, EC 2.3.1.20) catalyzes the acyl-CoA-dependent acylation of sn-1,2-diacylglycerol to form triacylglycerol (TAG). This enzyme appears to exert a substantial effect on the flow of carbon into TAG and might represent a bottleneck in seed oil production. Previously, several Brassica napus DGAT1 (BnaDGAT1) variants were generated via directed evolution, which resulted in increased oil content when the variants were produced recombinantly in the yeast (Saccharomyces cerevisiae) strain H1246. To explore the underlying causes of increased TAG production in the yeasts expressing BnaDGAT1 variants, the enzyme activity and polypeptide accumulation of different BnaDGAT1 variants were examined using yeast (strain H1246) microsomal fractions. Our results show that increased TAG accumulation was due to increased enzyme activity and/or polypeptide production. It is also interesting to note that one variant (L441P) was found to display both increased enzyme activity and possible reduced substrate inhibition. In addition, the amino acid residue substitutions shown to benefit BnaDGAT1 enzyme activity could be transferred to Camelina sativa DGAT1, suggesting that the information obtained from directed evolution of DGAT1 in one plant species could be transferred to other species. Thus, high performance forms of plant DGAT1 may be useful for increasing TAG production in oleaginous crops and microorganisms. Yang Xu: [email protected] SS.6: RUBY PARTICLES IN MUCILAGE (RUBY) is a putative glyoxal oxidase required for mucilage integrity and cell-cell adhesion in the seed coat epidermis of Arabidopsis thaliana Krešimir Šola1, Erin Gilchrist1, Marie-Christine Ralet3, Shawn Mansfield2, George Haughn1 1. Department of Botany, University of British Columbia, Vancouver BC, Canada 2. Department of Wood Science, University of British Columbia, Vancouver BC, Canada 3. INRA, Nantes, France When exposed to water, the seed coat epidermal cells of mature Arabidopsis thaliana seeds extrude mucilage composed mainly of unbranched pectin rhamnogalacturonan I (RG-I). Since mucilage is abundant and dispensable, we are using it as a model to study cell wall structure and function. The MUM2 gene encodes a β-galactosidase secreted into the mucilage. Loss-of-function mutations in MUM2 result in seed coat mucilage that cannot extrude, suggesting that modification of the mucilage by MUM2 is necessary to allow mucilage expansion during hydration. We have used genetic suppressor analysis to identify a new gene, RUBY PARTICLES IN MUCILAGE (RUBY), mutations in which suppress the lack of mum2 mucilage extrusion. In addition, seed coat epidermal cells separate from the seed coat in the ruby mutant. These phenotypes suggest that RUBY is needed for cohesion of both mucilage and middle lamellae. Analysis of cell wall carbohydrates released in the mutant suggests that RUBY functions by cross-linking a novel arabinogalactan-branched RG-I. RUBY encodes a glyoxal oxidase-related protein with a putative catalytic site that resembles those of previously studied glyoxal oxidases. Such enzymes generate hydrogen peroxide. We have been able to show that developing seeds of ruby mutants have a lower level of reactive oxygen species than wild type seeds consistent with the loss of glyoxal oxidase. We hypothesize that RUBY produces hydrogen peroxide that is needed to cross-link a branched RG-I thus strengthening both the mucilage and middle lamellae of seed coat epidermal cells. Krešimir Šola [email protected]

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Abiotic Stress I Wednesday July 5 1:30-3:00pm; FSC Room 1005 Chair: Annette Nassuth,, University of Guelph

AB1.1: Jasmonic acid and jasmonoyl-isoleucine are induced in response to mechanical wounding in the model lycophyte Selaginella moellendorffii Putri PRATIWI1*, Genta TANAKA1, Tomohiro TAKAHASHI1, Xiaonan XIE2, Koichi YONEYAMA2, Hideyuki MATSUURA1, Kosaku TAKAHASHI1 1. Research Faculty of Agriculture, Hokkaido University, Sapporo, Japan 2. Center for Bioscience Research and Education, Utsunomiya University, Utsunomiya, Japan Wounding is a common threat that affects plant growth and development. As a sessile organism, plants rarely escape damage from environmental stresses such as wind, rain, and herbivory attack thus plants evolved different mechanism to adapt to these circumstances. JA has been known to regulate a wide range of essential biological process in response to wound stress and is ubiquitously produced in vascular plants. However, instead of JA, OPDA probably play a main role in defense mechanism of bryophytes (a group of non-vascular plants). Here, we used Selaginella moellendorffii, a basal seedless vascular plant model to identify the presence of jasmonates. By using UPLC-MS/MS analysis, we demonstrated the first evidence of the presence of OPDA, JA, and JA-Ile in this plant. Therefore, the ability to synthesize JA in vascular plant and non-vascular plant might be one of the significant observations in the study of plant evolution. We also analyzed the functional activity of enzymes involved in JA biosynthesis and showed that SmAOS2, SmAOC1, SmOPR5, and SmJAR1 were actively involved in the production of OPDA, JA, and JA-Ile. The wound-induced temporal changes in OPDA, JA, and JA-Ile contents correlate tightly with the expression level of SmAOC1, SmOPR5, and SmJAR1. Furthermore, OPDA, JA, and JA-Ile inhibited the growth of S. moellendorffii. In conclusion, we propose the role of JA as a signaling molecule to regulate growth and response to wounding due to the acquisition of vascular system during plant evolution. Putri Pratiwi [email protected]

AB1.2: Nuclear localization and transactivation by Vitis CBF transcription factors are regulated by combinations of conserved amino acid domains Chevonne E. CARLOW1,2, J Trent FAULTLESS1, Christine LEE1,2, Mahbuba SIDDIQUA1,2, Alison EDGE1, Annette NASSUTH1 1. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON 2. Current Address: Ontario Ministry of Agriculture, Food and Rural Affairs, Vineland Station, ON 3. Current Address: Department of Plant Agriculture, University of Guelph, Guelph, ON The highly conserved CBF pathway is crucial in the regulation of plant responses to low temperatures. Extensive analysis of Arabidopsis CBF proteins revealed that their functions rely on several conserved amino acid domains although the exact function of each domain is disputed. The question was what functions similar domains have in CBFs from other, overwintering woody plants such as Vitis, which likely have a more involved regulation than the model plant Arabidopsis. A total of seven CBF genes were cloned and sequenced from V. riparia and the less frost tolerant V. vinifera. The deduced species-specific amino acid sequences differ in only a few amino acids, mostly in non-conserved regions. Amino acid sequence comparison and phylogenetic analysis showed two distinct groups of Vitis CBFs. One group contains CBF1, CBF2, CBF3 and CBF8 and the other group contains CBF4, CBF5 and CBF6. Transient transactivation assays showed that all Vitis CBFs except CBF5 activate via a CRT or DRE promoter element, whereby Vitis CBF3 and 4 prefer a CRT element. The hydrophobic domains in the C-terminal end of VrCBF6 were shown to be important for how well it activates. The putative nuclear localization domain of Vitis CBF1 was shown to be sufficient for nuclear localization, in contrast to previous reports for AtCBF1, and also important for transactivation. Annette Nassuth [email protected]

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AB1.3: Identifying the physiological and ultrastructural phenotypes responsible for chilling stress tolerance in corn. Tawhidur RAHMAN1 Valsala GOWRIBAI1, Perumal VIJAYAN1, Prakash VENGLAT1, Jitao ZOU2, Karen TANINO1 1Department of Plant Sciences, College of Agriculture and Bioresources, Univ. of Saskatchewan, Saskatoon, SK, Canada S7N 5A8 2National Reserch Council - Plant Biotechnology Institute (NRC-PBI), Saskatoon, SK, Canada S7N 0W9 Corn (Zea mays L.) is the largest crop grown globally. The crop is sensitive to chilling and frost, thus limiting its production in the colder climate regions in Canada. Currently, the production of corn is limited to just 0.3% of the arable land in Saskatchewan. Therefore, developing chilling and frost resistant corn varieties through breeding can contribute significantly to the agricultural economy of Saskatchewan. In our lab, we initiated an extensive stress physiological study in corn using a number cultivated genotypes available through Dow-DuPont-Pioneer. Our initial experimental data identified one cold sensitive (line:848) and one resistant (line: 884). When 20 seedlings of 848 and 884 lines were grown to the V2 stage and exposed to lethal chilling stress (day/night- 50C/20C; for 10 days) and allowed to recover (230C; 19 days), all of the 884 seedlings survived the chilling stress while none of of 848 seedlings survived. The water retention abilities of the excised V2 leaves of 884 were higher under controlled and stressed conditions compared to 848 plants. Acclimated and non-acclimated 884 seedlings also showed higher maximum photosynthetic efficiency than 848 seedlings under chilling stress. Chloroplastic phosphatidylglycerol fatty acid profiling data identified higher relative decrease of trans-16:1∆3 in the 884 line under chilling stress compared to the 848 line. We also monitored diurnal changes in leaf starch accumulation, which is a sensitive indicator of stress response of sugar translocation and storage processes, in these two lines. The data are under analysis and will be presented in the meeting. Collectively, our study aims to pinpoint the ultrastructural and key physiological parameters underlying chilling stress tolerance in corn. Tawhidur Rahman; email – [email protected]/[email protected]

AB1.4: Changes of the Grape Berry (Vitis vinifera L.) Cuticle during Fruit Development in Response to Water Deficit Stress. Nicolas DIMOPOULOS1*, Darren C.J. WONG1, Tegan HASLAM2, Changzheng SONG1 Rodrigo LOPEZ GUTIERREZ1, Ljerka KUNST2, Simone Diego CASTELLARIN1 1. Wine Research Centre, Faculty of Land and Food Systems, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 2. Department of Botany, Faculty of Science, University of British Columbia, Vancouver, BC V6T 1Z4, Canada In grapes, the cuticle protects the berry from water loss, which may in turn affect the final berry size at ripening. Water deficit (WD) is a common stress in vineyards, but little is known about how the berry cuticle is modified in response to this stress. Our previous work has shown that the expression of some cuticle genes is modulated in grape berries exposed to water deficit stress. Furthermore, differential expression and gene regulatory network analyses of grapevine RNA-seq datasets have indicated that the cuticular wax biosynthetic pathway is differentially regulated during berry development and in response to environmental stresses, including water deficit. Thus, we hypothesize that in response to WD stress, the developing grape berry will modify the composition of cuticular waxes on the berry surface to reduce the rate of water loss. We performed experiments in 2015 and 2016 to characterize changes in rate of water transpiration (drying experiment), in cuticular wax composition (GC-MS), and in biosynthetic gene expression (RT-qPCR) of Merlot berries developing under prolonged WD stress. Though the rate of water transpiration does not change, we observed changes in wax composition in our 2015 experiment. We expect to see reproducible results in the wax compositional and gene expression data we are currently analyzing from 2016. We expect significant changes in wax ester content and concurrent upregulation of the wax ester branch of the cuticular wax biosynthetic pathway starting at veraison (onset of ripening). Nicolas Dimopoulos [email protected]

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AB1.5: Can condensed tannins act as in vivo antioxidants and protect poplar against oxidative stress? Gerry GOURLAY*1, Barbara HAWKINS1, and C. Peter CONSTABEL1 1. Department of Biology, Forest Biology, University of Victoria, Victoria, BC Condensed tannins are polyphenolic compounds synthesized from the flavonoid pathway and the most abundant plant secondary metabolites. They accumulate to high concentrations in trees and are often considered to be plant defense compounds, but may have additional functions. In poplar, tannin biosynthesis is stimulated by high light intensity and nitrogen deficiency. Condensed tannins can act as potent in vitro antioxidants, but it is not known if they perform this function in planta. This project tests the hypothesis that condensed tannins can act as in vivo antioxidants and help to protect poplar leaves against oxidative stress. Transgenic poplar saplings engineered to accumulate high concentrations of condensed tannin were challenged with two stresses that cause accumulation of reactive oxygen species (ROS): treatment with the herbicide methyl viologen (Paraquat), and drought. Methyl viologen was applied to detached leaves via their petioles in the light, and chlorophyll fluorescence measured. After 24h of methyl viologen treatment, the high-tannin leaves retained significantly greater maximum quantum efficiency of photosystem II photochemistry (Fv/Fm) than control plants, indicating reduced photosystem damage. Drought stress was imposed by reducing water availability to saplings for three weeks. Chlorophyll fluorescence measurements showed that high-tannin transgenic poplar lines retained greater photosystem II operating efficiency (Fq’/Fm’, φPSII, or ΔF/Fm′) compared to control plants. These results suggest that condensed tannins help to protect against oxidative damage caused by these abiotic stressors. We speculate that tannins may also be important for tolerance to other abiotic stresses that generate ROS, such as UV-B or heat stress. Gerry Gourlay [email protected]

AB1.6: Effects of temperature, UVB radiation and watering regime on aerobic methane emissions during vegetative stages of pea plants Awatif M. ABDULMAJEED1*, Mirwais M. QADERI1,2 1. Department of Biology, Dalhousie University, Halifax, Nova Scotia B3H 4R2, Canada 2. Department of Biology, Mount Saint Vincent University, Halifax, Nova Scotia B3M 2J6, Canada Many studies have investigated the effects of single or double environmental factors on aerobic methane (CH4) emissions from plants, but the impacts of multiple factors have rarely been considered. We determined the effects of temperature, UVB radiation, and watering regime on CH4 emissions and other physiological parameters during three vegetative stages of pea plants (Pisum sativum L. cv. 237J Sundance) by growing them under controlled conditions: two temperature regimes (22/18oC and 28/24oC), two UVB levels [0 and 5 kJ m−2 d−1] and two watering regimes (field capacity and wilting point). Measurements were taken on 10-, 20-, and 30-day-old plants. Higher temperature, UVB5, and water stress increased CH4 emissions, which were highest at the earlier stages of plant growth. All three factors increased transpiration (E). Both higher temperatures and UVB5 increased stomatal conductance (gs), but only higher temperature decreased water use efficiency. Net CO2 assimilation, gs, and E. were highest in 30-day-old plants. Furthermore, higher temperature decreased the effective quantum yield of photosystem II ΦPSII and non- photochemical quenching, but increased photochemical quenching (qP), whereas water stress decreased ΦPSII, maximum quantum yield of PSII (Fv/Fm), and qP. Ten-day-old had highest ϕPSII, Fv/Fm, and qP, whereas 30-day-old plant had highest qNP. In addition, UVB5 decreases nitrogen balance index (NBI), but increase flavonoids. Ten-day-old plant had highest NBI and total chlorophyll, but had lowest flavonoid content. We conclude that temperature has the greatest influence on plant growth, and stress factors increase aerobic CH4 emission, which decreases with plant age. [email protected]

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Nutrients and Metabolism Wednesday July 5 1:30-3:00pm; FSC Room 1221 Chair: Lee Kalcsits, Washington State University NM.1: Advances and challenges for better managing nutrients in vegetable cropping systems Kate A. Congreves Department of Plant Sciences, University of Saskatchewan, Saskatoon, SK Traditional crop production research has focused heavily on agronomic and economic benefits of N fertilizer applications with little regard for the environment and nutrient use efficiency. But, now more than ever, producers and society at large recognize that sustainable food production requires management practices that balance the use of N for crop production while minimizing its environmental impact. Vegetable production presents a significant challenge for N management because many crops require large quantities of N fertilizer (i.e., 100 to >300 kg N ha-1) to maximize yields and profits - yet vegetable systems tend to be highly vulnerable for N loss, resulting in negative impacts to both the producer’s “bottom line” and the environment. Thus, improving crop N use in vegetable cropping systems may have economic and environmental benefits. Two management strategies aimed at reducing N loss without risking yields and production include: (i) reducing excessive N fertilizer applications during the growing season and (ii) implementing a cropping system which sequesters N during the non-growing season to prevent N loss. This talk will delve into the opportunities and tribulations towards developing better N management practices for vegetable cropping systems. [email protected] NM.2: On the path towards developing plants that express a functional nitrogenase Perrin H BEATTY1, Julia L WONG1,2 and Allen G GOOD1 1. Department of Biological Sciences, University of Alberta, Edmonton, AB, Canada 2. School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, Australia Nitrogen (N), an essential element for plants, is acquired as fixed-N from the soil and from nitrogen fixing bacteria. For crop plants, the level of available fixed-nitrogen directly influences biomass and grain yield. N tends to be scarce in poor soils and over-abundant in rich soils, resulting either low yielding plants or plants that can only take up approximately 50% of the supplied N, leading to polluted aquatic and atmospheric ecosystems. One solution aimed at both increasing yields and reducing N pollution is to engineer nitrogen-fixing crop plants with the genes required to express a functional nitrogenase. Can the ability to fix N2 be successfully introduced into crop plants? The long-term goal of this project is to synthesize biologically active N within plant mitochondria (Mt), so that N supply can be tightly coupled to available carbon within the plant, and ultimately growth and yield. Mt-tagged nifH genes from two diazotrophic bacteria plus nifM from Azotobacter vinelandii were transiently introduced into tobacco leaves using Agrobacterium tumefaciens infiltration. The plant expressed NifH protein was purified anaerobically from the leaves as a crude mitochondrial extract and used in a functional acetylene reduction assay complemented with purified NifDK complex. The results of these experiments will be discussed, focussing on the challenges faced in expressing a functional NifH protein in planta. Given the economic importance of fixed N2 and the significant environmental pollution caused by excessive N, it is time to refocus our efforts and resources on making crop plants that can fix nitrogen. Perrin Beatty [email protected]

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NM.3: Yield response of wheat, pea and canola to micronutrient fertilization in contrasting prairie soils

Noabur RAHMAN1* and Jeff SCHOENAU1 1. Department of Soil Science, University of Saskatchewan, Saskatoon, Canada Assessing the need for micronutrient fertilization and factors influencing the likelihood of crop response is a concern in the development of crop nutrition plans for growers on the prairies. It is not well known what extent of deficiencies of Cu, Zn and B conclusively exist across the prairies, and the degree to which wheat, pea, and canola crops may respond to fertilization with these micronutrients. The objective of this research is to evaluate the impact of micronutrient fertilization on yield of crops grown in contrasting prairie soils under polyhouse conditions. Five different soils with a range of micronutrient fertility from deficient to sufficient according to standard soil test method are used to grow wheat, pea and canola crops. Important parameters including micronutrient fertilizer form, application method, and crop yield responses are examined. The grain yield of wheat was significantly increased by Cu fertilization in two of the five soils. Initial Cu fertility was below critical level in one soil (Ukalta), while the other one considered as sufficient (Sceptre) based on soil-test result interpretations. No significant response of peas to Zn fertilization was recorded on any of the five soils. Canola significantly responded to B fertilization on one soil (Whitefox). The soil was critically deficient in B and only soil application was effective in correcting the problems for Canola production. Noabur Rahman, [email protected] NM.4: A curculin-like lectin interacts with a high mannose glycoform of the purple acid phosphatase AtPAP26 in cell walls of phosphate-starved Arabidopsis thaliana Mina GHAHREMANI1*, Erin ANDERSON3, Robert MULLEN3, Yi-Min SHE2, William C. PLAXTON1

1. Department of Biology, Queen’s University, Kingston, ON 2. Centre for Biologics Evaluation, Health Canada, Ottawa, ON 3. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON Purple acid phosphatases (PAPs) function in the acquisition and recycling of Pi, a crucial but environmentally limiting macronutrient for plant growth. Among 29 predicted Arabidopsis PAPs, AtPAP26 (At5g34850) functions as the principal vacuolar (V), cell wall (CW), and root secreted PAP isozyme upregulated during Pi-deprivation. A pair of differentially glycosylated AtPAP26 ‘glycoforms’ were resolved from CW extracts of Pi-deprived Arabidopsis suspension cells during lectin-affinity chromatography. High resolution LC-MS/MS demonstrated that glycan structures of purified AtPAP26-CW1 and -CW2 at Asn103 and Asn365 were virtually identical. However, AtPAP26’s Asn422 glycosylation site was extensively modified in AtPAP26-CW2 by the addition of up to seven terminal mannose residues (to N-acetylglucosamine) to form a high mannose glycan. A 55-kDa protein co-purified with AtPAP26-CW2 and was identified by LC MS/MS as an Arabidopsis curculin-like (mannose-binding) lectin (At1g78850). Parallel qPCR and anti-curculin immunoblotting indicated that curculin is significantly upregulated at both the transcript and protein level in Pi-starved Arabidopsis. Reciprocal far-western immunoblotting demonstrated a specific interaction between purified curculin and AtPAP26-CW2, but not AtPAP26-CW1. Bifluorescence-complementation and CLSM revealed an in vivo interaction between AtPAP26 and curculin within the vacuole of the Pi-deficient suspension cells. AtPAP26-CW2’s acid phosphatase activity and thermal stability were enhanced after it was preincubated with curculin. Research is in progress to assess the phenotype of Arabidopsis T-DNA insertional curculin ‘knockout’ plants. We hypothesize that curculin plays a key role during Pi deprivation through its interaction with mannose-rich oligosaccharide groups of AtPAP26-CW2, and consequent impact on AtPAP26-CW2 function and stability. Mina Ghahremani: [email protected]

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NM.5: Mosses get the munchies Barbara HAWKINS, Emily MAY, Samantha ROBBINS Centre for Forest Biology, University of Victoria, Victoria, BC Mosses are found in every terrestrial ecosystem on earth. British Columbia has the highest diversity of mosses in Canada and mosses can be found in widely contrasting environments. Lacking a vascular system and roots, mosses must take up nutrients through leaf and stem surfaces; but as found for vascular plants, nitrogen (N) is the nutrient required in the greatest amount. Little is known about N uptake by mosses, thus the objective of this study was to measure rates of ammonium and nitrate uptake by a variety of moss species from contrasting habitats. Net fluxes of ammonium, nitrate and protons were measured on leaf surfaces of 25 moss species from high and low rainfall sites on Vancouver Island using a microelectrode ion flux measurement (MIFE) system. Most moss species exhibited efflux of protons from leaves and had higher rates of ammonium uptake than nitrate uptake. Rates of N uptake in leaves of many moss species were similar to N uptake rates previously measured in conifer roots. Moss species with the highest rates of uptake of both ammonium and nitrate were from dry, rocky or epiphytic habitats. Despite their small stature, mosses can sequester significant quantities of available N on forest sites. Barbara Hawkins ([email protected])

NM.6: Carbonic anhydrase activity is related to mesophyll conductance in black cottonwood genotypes Mina Momayyezi1*, Robert D. Guy1

1. Department of Forest and Conservation Sciences, Faculty of Forestry, University of British Columbia Ecophysiological traits of plant species, distributed over geographically wide ranges, vary with environmental gradients. In black cottonwood (Populus trichocarpa Torr. & Gray), clinal variation in photosynthetic rate (An) is partly supported by greater stomatal conductance (gs) in northern genotypes. Gas exchange analysis (A:Ci curves), chlorophyll fluorescence and on-line stable carbon isotope discrimination were used to measure mesophyll conductance (gm) in this species. Across 12 native genotypes of black cottonwood, gm increased with latitude of origin in parallel with An. This pattern may be related to clinal variation in structural and/or physiological

components affecting gm. Carbonic anhydrase (CA) activity facilitates the CO2 HCO3− equilibrium, and can

thereby influence diffusion through the liquid phase, but definitive experimental evidence for a significant effect of CA on gm or photosynthesis has been lacking. Petiolar feeding with acetazolamide (a CA inhibitor), simultaneously reduced CA activity, gm, gs, chloroplast CO2 concentration (Cc) and An at ambient CO2 (400 µmol mol-1). The effect on An was reversible at saturating CO2, confirming that acetazolamide had no direct effect on photosynthesis other than causing a reduction in gm. Regardless of expression on either a per unit leaf mass or area basis, three representative northern genotypes had greater than two fold higher CA activity than three representative southern genotypes, consistent with their almost two fold higher gm. It is suggested that greater An in northern genotypes is supported by their higher gm compared to southern genotypes, which, in turn, is strongly associated with their greater CA activity. [email protected]

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Sexual Reproduction Wednesday July 5 3:20-4:50pm; FSC Room 1003 Chair: Hong Wang, University of Saskatchewan SR.1: Arabidopsis ICK/KRP cyclin-dependent kinase inhibitors are critical for ensuring the development of one megaspore mother cell and one functional megaspore per ovule Ling CAO1,2 *, Sheng WANG1, Prakash VENGLAT3, Lihua ZHAO4, Yan CHENG1,2, Shengjian YE1, Yuan QIN4, Raju DATLA3, Yongming ZHOU2 and Hong WANG1 1Dept. of Biochemistry, University of Saskatchewan, Saskatoon SK, S7N 5E5, Canada; 2National Key Laboratory of Crop Genetic Improvement, Huazhong Agricultural University, Wuhan 430070, China; 3National Research Council Canada, Saskatoon, SK, S7N 0W9, Canada; 4Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Fujian Agriculture and Forestry University, Fuzhou, Fujian 350002, China In most plants, the crucial and initial step for the transition from somatic to female germline development is the differentiation of megaspore mother cell (MMC). In this process, a single subepidermal cell in the nucellus of a young ovule differentiates and enlarges to become an MMC. The MMC produces multiple megaspores through meiosis, one of which survives to become the functional megaspore (FM). The single embryo sac formed from the FM is subsequently fertilized producing one embryo. Little is known regarding how the MMC and FM are selected. The ICK/KRPs (interactor/inhibitor of cyclin-dependent kinase (CDK)/Kip-related proteins) are plant CDK inhibitors and cell cycle regulators. Here we report that in the ovules of Arabidopsis mutant with all seven ICK/KRP genes inactivated, there were multiple MMCs, multiple FMs and multiple embryo sacs, which could be fertilized to form twin embryos with separate endosperms. Twin seedlings were observed in about 2% seeds. Also, in the mutant ovules the number and position of surviving megaspores among the megaspores from one MMC were not fixed, but variable. The absence of or much weaker phenotypes in lower orders of mutants and complementation of the septuple mutant by ICK4 or ICK7 indicate that multiple ICK/KRPs function redundantly to restrict the differentiation of more than one MMC and also likely to suppress the formation of multiple FMs, which is critical to ensure the development of one embryo sac and one embryo per ovule. Ling CAO [email protected] SR.2: The BRASSIKIN (BKN) pseudokinases modulate receptor complex and family-specific mate acceptance

Hyun Kyung Lee*1, Daphne R. Goring1

1. Department of Cell and Systems Biology. University of Toronto, ON, Canada

Plant have developed ways to recognize compatible pollen grains as providing nutrients to the wrong partner is detrimental. The Brassicaceae family of plants employ tight regulation in accepting pollen by choosing to deliver cellular components from the stigma to allow for pollen grain hydration and pollen tube growth. Physiological processes that facilitate such processes are well known; however, the initial upstream signalling components are yet to be elucidated. The BKNs are candidate stigma signalling proteins that were identified in the Goring lab where transgenic knockdown lines displayed reduced pollen hydration and seed production. BKNs are highly conserved pseudokinases within the Brassicaceae and likely to work in conjunction with other signalling proteins. We hypothesized that BKNs mediate signal transduction by forming a receptor complex with receptor-like kinases (RLKs) to facilitate pollen hydration. To elucidate BKNs’ putative interactors, Y2H screening was conducted in pairwise interactions with RLKs that were chosen based on two stigma microarray datasets and one stigmatic papilla RNA-seq dataset. Putative interactors were identified and found to cluster to two distinct RLK sub-families. Bimolecular fluorescence complementation assay was conducted to confirm protein interactions and their subcellular localization in planta. Most importantly, to test for the biological relevance of these interactors, CRISPR/Cas9 system was used to delete a cluster of four candidate RLK genes, and these knockout plants are currently being characterized for altered pollen-stigma interactions. Overall, this project will provide insights into

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the potential roles of this RLK subfamily in the stigma pathway regulating mate recognition and initiating pollen hydration.

Hyun Kyung Lee, [email protected]

SR.3: Understanding role of Phospholipase D1 during pollen-pistil interactions in canola Sabine SCANDOLA1*, Marcus A. SAMUEL1 1. Department of Biological Sciences, University of Calgary, 2500 University Drive, NW Calgary, AB, T2N 1N4, Canada Self-incompatibility (SI) is a genetic mechanism in hermaphroditic flowers that prevents inbreeding by rejection of self-pollen while allowing cross or genetically different pollen to germinate on the stigma to successfully fertilize the ovules. In Brassicaceae (A. thaliana, canola, broccoli, cabbage, kale), this process is triggered by activation of the receptor kinase in the papillary cells of the stigma following binding of haplotype-specific ligands produced by the pollen. Once activated, this phospho-relay converges on compatibility factors, which are immediately targeted for degradation, resulting in pollen rejection response. Through proteomics approaches, we have identified phospholipase D1 (PLD1) as one of the proteins that is targeted for degradation through this process. Loss of PLD1 leads to a reduction in pollen attachment and pollen tube penetration following compatible pollination, while overexpression of PLD1 in the stigmas is breaking down SI response. Localization studies and visualization of multivesicular bodies (MVBs) in the papillary cells through electron microscopy, allowed us to characterize PLD1 function at a whole stigma as well as at a subcellular level. Papillary cells of the PLD1 overexpression lines were found to be promoting significantly more fusion of MVBs to the plasma membrane following pollination than the control lines. We predict that phosphatidic acid produced as a result of PLD1 activity is involved in modification of membrane curvatures which favors membranes fusion, allowing exocytosis of compartments (MVBs) containing compatibility factors. Sabine Scandola [email protected]

SR.4: Discovery of the regulatory hierarchy of the unicellular zygospore development using the Chlamydomonas model system

Thamali KARIYAWASAM1*, Sunjoo JOO 1, Ran Ha HONG1, Deepak TOOR1, Evan CRONMILLER1, Kyungchul NOHH2, Tuya WULAN2, and Jae-Hyeok LEE 1 1. Department of Botany, University of British Columbia, Vancouver, Canada, 2. Department of Biology, Washington University in St. Louis, St. Louis, USA

Gametic fertilization culminates in embryogenesis establishing major organ systems in plants, whereas it results in the zygospore differentiation in most green algal species including a unicellular green alga, Chlamydomonas reinhardtii. Therefore, the zygospore differentiation of Chlamydomonas can serve as a model for the ancestral developmental program that gave rise to the plant embryogenesis. The Chlamydomonas sexual life cycle consists of two genetically defined sexual gametes, plus and minus, whose syngamy is sufficient to execute the whole zygospore program by the heterodimerization of two TALE-class homeobox proteins, Gsm1 and Gsp1, homologous to the KNOX/BELL heterodimerization involved in shoot apical meristem and organ development. However, we know little about how the Gsm1/Gsp1-dependentzygospore program regulates the entire differentiation program. To identify molecular players involved in the zygospore program, we generated ~ 15,000 insertional mutants using an engineered strain capable of selfing to produce homozygous 2N zygotes, which allow direct screening for mutants defective in the zygospore differentiation. So far, genetic analysis has confirmed 23 mutants that are divided into gametic defect and zygotic defect groups. Using the pleiotropy of their cellular defects, we propose a genetic hierarchy model for the zygospore differentiation. Identification of their causative mutations will be presented to illustrate the complexity of the developmental regulatory network evolved for the sexual life cycle.

[email protected]

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SR.5: From profiles to phenotype: epigenome analysis reveals gender-specific methylation of a gene in the sex-determining region of Populus balsamifera Katharina BRAEUTIGAM1, Raju SOOLANAYAKANAHALLY2, Marc CHAMPIGNY3, Shawn D. MANSFIELD4, Carl DOUGLAS5, Malcolm M. CAMPBELL6, Quentin CRONK5 1. Department of Biology, University of Toronto Mississauga, Mississauga, ON, Canada 2. Saskatoon Research and Development Centre, Agriculture and Agrifood Canada, Saskatoon, SK, Canada 3. Department of Biological Sciences, University of Toronto Scarborough, Toronto, ON, Canada 4. Department of Wood Science, University of British Columbia, Vancouver, BC, Canada 5. Department of Botany, University of British Columbia, Vancouver, BC, Canada 6. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON, Canada In addition to the genetic information, epigenetic factors have been widely implicated in shaping cell function and ultimately phenotype. Epigenetic factors can modify chromatin structure, modulate gene expression and thus contribute to genome interpretation and function. Here, we investigate genome-wide DNA methylation profiles in relation to gender in Populus balsamifera, a tree with a wide natural distribution and economic importance in Canada. Sexual reproduction is key to a vast majority of organisms on earth, and the molecular mechanisms contributing to sex-determination have been relatively well-characterized in a number of model organisms such as C. elegans, Drosophila, or mouse. Similar to animals, some angiosperms such a trees in the genus Populus have separate male and female individuals (dioecy). However, much remains to be learned about the molecular mechanisms shaping dioecy in angiosperms. Genome-wide DNA methylation profiles were obtained from multiple male and female balsam poplar trees. Targeted as well as non-targeted analyses (statistical learning) revealed a strong sex-specific differential methylation in PbRR9, a two-component response regulator gene. Intriguingly, this gene is also located in the recently identified genetic sex-determining region of this species. Ongoing analyses focus on a detailed mechanistic understanding of sex determination in P. balsamifera, and findings might also have potential applications to breeding in dioceous plants. [email protected]

SR.6: Furtive Flowers: Phenology, shoot development, and organization of pistillate Arceuthobium americanum

Dylan J. Ziegler1 & Cynthia M. Ross Friedman1 1. Thompson Rivers University, Kamloops, BC Dwarf mistletoes are dioecious parasitic angiosperms having prolonged life cycles lasting six years, culminating with explosive discharge of the single seed from the fruit. Arceuthobium americanum, the lodgepole pine dwarf mistletoe, infects lodgepole pine (Pinus contorta) in western North America, compromising the forest economy. While our understanding of the biology underlying the dwarf mistletoe-lodgepole pine interaction is becoming clearer, we still know little about the female’s life cycle, and nothing about characteristic branching patterns. Here, we used environmental scanning electron microscopy to demark phenological waypoints throughout the pistillate plant’s development to identify branching and growth patterns. We found highly reduced flowers appear no later than two years following aerial shoot development, with female flowers appearing and persisting for over two years. Development of the pistillate plant, including initiation of floral growth, has not been explained in detail. As successive crops of female flowers emerged every year, up to three generations of flowers/fruit could be found on a single shoot in late summer; we used these three generations to delineate specific developmental stages. Vegetative shoots initiated growth at any time within the growing season and assumed either a terminal position or adopted sympodial branching, but were never whorled/adventitious. Floral branches could initiate adventitiously from older nodes in a whorled pattern, and could house flowers/fruits of any generation. Vegetative and floral units were structurally homogeneous, suggesting shared developmental pathways. In identifying these phenological stages of A. americanum, control strategies to minimize lumber damage can accurately target vulnerable stages in the plant’s development. Dylan Ziegler [email protected]

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Specialized Metabolism I (Anatomy and Defence) Wednesday July 5 3:20-4:50pm; FSC Room 1221 Chair: Peter Constabel, University of Victoria SM1.1: Localization and Ecological Functions of Condensed Tannins in Poplar Roots C. Peter CONSTABEL1, Rebecca WESTLEY1, and Barbara HAWKINS1 1. Centre for Forest Biology and Department of Biology, University of Victoria, Victoria, BC The condensed tannins, also called proanthocyanidins, are the most widely distributed secondary plant metabolites. They are especially abundant in woody plants, where they can accumulate to 25 % of leaf dry weight. Condensed tannins (CTs) in leaves are implicated in plant defense, especially against vertebrate herbivores, but also play additional roles. They are also very abundant in tree roots; surprisingly, very little is known about their functional significance in below-ground organs. To address this knowledge gap and to develop hypotheses regarding the role of CTs in roots, we mapped the accumulation of CTs in roots of poplar saplings. CT concentrations were greatest in root tips and in young white roots, and diminished with age and in the brown portions of the root system. Using a 4-dimethylaminocinnamaldehyde (DMACA), we localized the CTs to specific cell types. In root tips, tannins accumulated in epidermal layers and the root cap. Slightly older white roots also accumulated CTs in the epidermis, as well as in randomly distributed cells within the cortex. To investigate a possible spatial correlation of CTs with nutrient uptake, we used a microelectrode ion flux measurement (MIFE) system to measure rates of nitrogen fluxes along the root axis. Nitrate and ammonium uptake was maximal in the first 5 mm of the tip, but did not correlate with CT distribution. Based on their distinct localization, we hypothesize that root CTs are important in below-ground defense. This idea will be tested using RNAi transgenic poplars with reduced CT content. Peter Constabel [email protected] SM1.2: Transcriptome reprograming during resin duct biogenesis in white spruce (Picea glauca) cambium cells Jose M. CELEDON, Angela CHIANG, Macaire M.S. YUEN and Jörg BOHLMANN Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T 1Z4, Canada Conifers have multiple chemical and physical defenses that are key to their adaptability and survival. Among them, terpene-rich oleoresin is thought to play a central role in conifer’s multi-layered defense systems against insects and pathogens. Under normal, non-stress conditions, oleoresin is constitutively produced and stored in cortical resin ducts (CRD) forming a protective layer in stem bark tissue. Upon insect attack, or methyl jasmonate (MeJa) treatment, stems of several conifer species including Picea glauca, form a second layer of resin ducts, called traumatic resin ducts (TRD), that arises de novo from vascular cambium cells. Much of our knowledge on conifer terpenoid defenses is based on characterized terpene synthases (TPS) and cytochrome P450s (P450) that are highly and specifically expressed in the cells lining CRDs and TRDs. However, knowledge on the biogenesis of resin ducts remains limited and represents a major gap in our understanding of conifer terpenoid defenses. To gain insight into resin duct development, we sequenced the transcriptome of cambium cells microdissected from P. glauca trees treated with MeJa and followed its changes over time. We identified a group of MYB and WRKY transcription factors having significantly increased expression in cambium cells in response to MeJa suggesting they may participate in TRD formation. We also observed up-regulation of known TPS and P450 transcripts at 24 h after treatment suggesting an early activation of terpenoid biosynthesis. Knowledge of regulatory genes and networks involved in TRD biogenesis will significantly improve our understanding of terpenoid-based defenses and development of secretory structures in conifers. Jose M. Celedon [email protected]

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SM1.3: Transcriptome reprograming during resin duct biogenesis in white spruce (Picea glauca) cambium cells Jose M. CELEDON, Angela CHIANG, Macaire M.S. YUEN and Jörg BOHLMANN Michael Smith Laboratories, University of British Columbia, Vancouver, BC V6T 1Z4, Canada Conifers have multiple chemical and physical defenses that are key to their adaptability and survival. Among them, terpene-rich oleoresin is thought to play a central role in conifer’s multi-layered defense systems against insects and pathogens. Under normal, non-stress conditions, oleoresin is constitutively produced and stored in cortical resin ducts (CRD) forming a protective layer in stem bark tissue. Upon insect attack, or methyl jasmonate (MeJa) treatment, stems of several conifer species including Picea glauca, form a second layer of resin ducts, called traumatic resin ducts (TRD), that arises de novo from vascular cambium cells. Much of our knowledge on conifer terpenoid defenses is based on characterized terpene synthases (TPS) and cytochrome P450s (P450) that are highly and specifically expressed in the cells lining CRDs and TRDs. However, knowledge on the biogenesis of resin ducts remains limited and represents a major gap in our understanding of conifer terpenoid defenses. To gain insight into resin duct development, we sequenced the transcriptome of cambium cells microdissected from P. glauca trees treated with MeJa and followed its changes over time. We identified a group of MYB and WRKY transcription factors having significantly increased expression in cambium cells in response to MeJa suggesting they may participate in TRD formation. We also observed up-regulation of known TPS and P450 transcripts at 24 h after treatment suggesting an early activation of terpenoid biosynthesis. Knowledge of regulatory genes and networks involved in TRD biogenesis will significantly improve our understanding of terpenoid-based defenses and development of secretory structures in conifers. Jose M. Celedon [email protected] SM1.4: Lavandula x intermedia 3-carene synthase catalyzes pernyl diphosphate isomers (GPP and NPP) to produce distinct levels of monoterpenes in vitro Ayelign M. ADAL*, Soheil S. MAHMOUD University of British Columbia - Okanagan , Department of Biology, 1177 Research Road, Kelowna, BC Canada V1V 1V7 Lavandula essential oils that are composed of mainly monoterpenes play several economical and ecological roles. 3-Carene - one of the monoterpenes presented in Lavandula oils - has antimicrobial and insecticidal properties from different plants. However, the biosynthesis and spatio-temporal regulations of 3-carene remain unknown in Lavandula. Hence, in this study we isolated and functionally characterized 3-carene synthase (Li3CARS) from Lavandula x intermedia in vitro. We expressed the open reading frame excluding transit peptides in E. coli, and purified with Ni-NTA agarose affinity chromatography for in vitro assay. The recombinant partially purified Li3CARS converted geranyl diphosphate (GPP) into 3-carene as the major product and other minor monoterpenes. Recombinant Li3CARS also catalyzed neryl diphosphate (NPP), the cis-isomer of GPP, to produce multiple products with a small quantity of 3-carene. Li3CARS has higher affinity and catalytic efficiency for GPP (Km = 3.69 ± 1.17 µM, kcat/Km = 0.56 µM-1s-1) than NPP (Km = 14.3 ± 2.56, kcat/Km = 0.044 µM-1s-1) to produce 3-carene. The affinity of different substrates by Li3CARS to produce distinct monoterpenes in vitro suggests that substrate availability may partly control monoterpene metabolism in plant cells. Li3CARS was differentially expressed, and was highly abundant in leaves as compared to flower tissues, which positively correlated with 3-carene accumulation. Both Li3CARS transcript and 3-carene were also up-regulated with exogenous methyl jasmonate, suggesting that Li3CARS may function as a defensive compound in Lavandula. Ayelign Adal [email protected]

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SM1.5: Natural rubber and sesquiterpene lactones co-accumulate in laticifer but their promoters show differential expression patterns in lettuce. Connor L. HODGINS1*, Elysabeth REAVELL-ROY1, Moonhyuk KWON1, Eun-Joo KWON1, Dae-Kyun RO1 1. Department of Biological Sciences, University of Calgary, Calgary, AB, Canada Lettuce (Lactuca sativa) accumulates two distinct terpenoid products, natural rubber (NR) and sesquiterpene lactones (STL), in specialized laticifer cells. Although the biosynthesis of NR and STLs has been partially elucidated, it remains unknown how the transcription of their metabolic genes, which share the precursor isopentenyl diphosphate (IPP), is regulated at the cellular level. To investigate the transcriptional regulation of the biosynthetic NR and STL genes, promoters of cis-Prenyltransferase Binding Protein isoform 2 (LsCBP2) and germacrene A synthase (LsGAS) were isolated from lettuce genomic DNA. Their cellular specific expression patterns were examined in transgenic lettuce plants by expressing ®-glucuronidase (GUS) transcriptionally fused to either the LsCBP2 or LsGAS promoter. Both transgenic lines showed specific GUS-staining patterns along the vasculature. However, fluorescent assays using isolated latex showed the latex from LsCBP2-GUS lines had three-orders of magnitude higher levels of GUS activity than those from LsGAS-GUS. Quantitative PCR data showed exclusive expression of LsCBP2 in latex while LsGAS showed higher expression in whole stem compared to latex. Sectioning of the GUS-stained transgenic lettuce plants demonstrated that the LsCBP2 promoter drives expression in latex, whereas the LsGAS promoter displayed strong expression in the parenchymal cells surrounding the laticifer. These results suggest that transcription of LsGAS and LsCBP occur in different cell-types in lettuce, even though their metabolic products, NR and STL, co-accumulate in laticifers. This metabolic compartmentalization helps to explain how copious amounts of two different terpenoids co-accumulate in a single cell-type and further indicates the presence STL-trafficking from parenchyma cells to laticifer cells. Connor Hodgins, [email protected]

SM1.6: Terpene synthases from Cannabis sativa Judith K. BOOTH1, Jonathan E. PAGE2.3, Jörg BOHLMANN1,3 1. Michael Smith Laboratories, University of British Columbia, Vancouver, BC 2. Anandia Laboratories, Lower Mall, Vancouver, BC 3. Botany Department, University of British Columbia, Vancouver, BC

Cannabis (Cannabis sativa) produces a terpene-rich resin in glandular trichomes, which are abundant on the surface of the female inflorescence. The resin is valued for its medicinal and psychoactive properties. Monoterpenes and sesquiterpenes are important components of cannabis resin, as they define some of the unique organoleptic properties. Terpenes may also influence the medicinal qualities of cannabis strains and varieties. Transcriptome analysis of trichomes of the cannabis varieties ‘Finola’ and ‘Purple Kush’ revealed sequences of all stages of terpene biosynthesis. Fifteen cannabis terpene synthases (CsTPS) were identified in subfamilies TPS-a and TPS-b. Functional characterization identified mono- and sesqui-TPS, whose products collectively comprise many of the terpenes reported in cannabis. Products include compounds such as myrcene, (E)-β-ocimene, (-)-limonene, (+)-α-pinene, β-caryophyllene, and α-humulene. Transcripts associated with terpene biosynthesis are highly expressed in trichomes compared to non-resin producing tissues. Knowledge of the CsTPS gene family may offer opportunities for selection and improvement of terpene profiles in cannabis. Judith Booth [email protected]

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Abiotic Stress II Wednesday July 5 3:20-5:05pm; FSC Room 1005 (7 talks) Chair: Simone Castellarin, University of British Columbia AB2.1: Revealing the response of fruit metabolism to drought in grapevine (Vitis vinifera L.); omics applications in applied physiology

Simone D. CASTELLARIN1 1. Wine Research Centre, University of British Columbia, Vancouver, BC Drought is considered one of the major threats for crops in the predicted future climatic scenarios. In fruit crops, drought can impact the accumulation of metabolites that determine fruit quality. Classified as drought tolerant, grapevine is often not irrigated or minimally irrigated to improve the berry composition. In these situations, water deficit can promote the synthesis of volatile organic compounds, carotenoids, and phenolics in the berry. Network-based analysis using multi-omics data provide a powerful tool for discovering links between and within the many layers of biological complexity, such as the coordinated regulation of genes and metabolic pathways that govern the grape berry response to drought. We aimed to apply this approach to deeply characterize the molecular and metabolite response to water deficit, to uncover the relationships between water-deficit responsive genes and the metabolite accumulation during berry development, and to identify key putative molecular regulators that underlay the metabolic response to water deficit. Large scale metabolite and transcript (RNA-Sequencing) analyses were applied to white and red grapes exposed to drought during the growing season. The integrated network and promoter analyses identified transcriptional regulatory modules that encompassed terpenoid or flavonoid genes, transcription factors, and enriched drought-responsive elements in the promoter regions of those genes as part of the grapes response to drought. This integrated approach shows that water deficit-regulated gene modules are strongly linked to key fruit-quality metabolites, and multiple signal transduction pathways may be critical to achieve a balance between the regulation of the stress-response and the berry ripening program. Simone D. Castellarin [email protected]

AB2.2: AITRs function as feed-forward regulators of ABA signaling and are involved in the regulation of abiotic stress tolerance in Arabidopsis

Hainan TIAN, Siyu CHEN, Wenting YANG, Tianya WANG, Kaijie ZHENG, Yating WANG, Yuxin CHENG, Na ZHANG, Shanda LIU, Dongqiu LI, Bao LIU, Shucai WANG* Key Laboratory of Molecular Epigenetics of MOE, Northeast Normal University, Changchun, Jilin, China Abiotic stresses are global problems that limit yield in crops and other plants. The plant hormone abscisic acid (ABA) plays a crucial role in regulating plant responses to abiotic stresses. ABA signaling is triggered by binding of ABA molecules to the PYR/PYL/RCAR receptors, which promotes the interaction of the receptors with PP2C protein phosphatases, resulting in de-repression of the SnRK2 protein kinases, sequentially the activation of downstream transcription factors, leading to the activation of ABA response genes. We found that ABA-induced transcription repressors (AITRs), a novel family of transcription factors play a feed-forward role in ABA signaling. RT-PCR results showed that the expression of all the six Arabidopsis AITR genes was induced by exogenously ABA, and their basel expression levels were decreased in the ABA biosynthesis mutant aba1-5. When recruited to the promoter region of a reporter gene by a fused DNA binding domain, all AITRs tested, including Arabidopsis, tomato, rice and soybean family members, inhibited reporter gene expression in transfected protoplasts. In Arabidopsis, aitr mutants showed reduced sensitivity to ABA and to stresses such as salt and drought. Quantitative RT-PCR analysis demonstrated that the ABA-induced response of PP2C and some PYR/PYL/RCAR genes was reduced in AITR5 transgenic plants, but increased in an aitr2 aitr5 aitr6 triple mutant. BLAST searches showed that AITRs are exclusively present in angiosperms. These results provide important new insights into the regulation of ABA signaling in plants, and such information may lead to the production of plants with enhanced resistance to abiotic stresses.

[email protected]

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AB2.3: Unraveling transcriptomic responses of an allopolyploid to abiotic stresses: homoeologous expression and alternative splicing patterns JOON SEON LEE1*, DAVID C. TACK2, KEITH L. ADAMS1 1. Department of Botany, University of British Columbia, Vancouver, BC Canada 2. Department of Biology, Penn State University, State College, PA USA Polyploidy is one of the major driving forces and a prevalent phenomenon in plant evolution. Understanding the abiotic stress response of the polyploids is crucial since many agriculturally important crops are polyploids. Homoeologous differential expression and differential splicing patterns under different abiotic stress conditions may explain how increased-size plant genomes respond and adapt to the abiotic stresses. In this study, we have conducted the most comprehensive transcriptome analysis of Brassica napus, a tetraploid derived from Brassica rapa (AT) and Brassica oleracea (CT), by ultra-high throughput transcriptome sequencing from 12 samples in four conditions. We found AT subgenome biased gene expression and CT subgenome biased alternative splicing patterns (quantitatively and qualitatively) in the homoeolog sets (27,360 pairs) in all conditions. Coding gene expression under abiotic stresses was negatively correlated with antisense transcripts (P<0.01). Cold stress made the greatest gene expression changes compared to the other conditions. These plants under cold stress and drought stress shared more alternative splicing patterns than heat stress. Alternative splicing variants were confirmed by PCR from randomly selected genes. We suggest that AT subgenome is more responsive to the abiotic stresses than CT subgenome, but CT subgenome could be strongly engaged with down-regulation mechanism via nonsense-mediated decay. Joon Seon Lee [email protected]

AB2.4: microRNA156 regulates drought tolerance strategies in Medicago Sativa at the transcriptomic, metabolomic and physiological levels Biruk A Feyissa*1, 2, Susanne Kohalmi1, Abdelali Hannoufa1, 2 1. University of Western Ontario, N6A 3K7, London, ON, Canada 2. Agriculture and Agri-Food Canada, N5V 4T3, London, ON, Canada

Developing alfalfa (Medicago sativa) cultivars that can withstand drought is critical for its sustainable production. Among hundreds of microRNAs discovered in plants, miR156 is highly conserved in plants, where it functions by downregulating a group of SQUAMOSA-PROMOTER BINDING PROTEIN-LIKE (SPL) transcription factors. In this study, we used three alfalfa genotypes with different levels of miR156 overexpression and genotypes with reduced SPL13 expression to study drought response at the phenotypic, physiological, metabolic and molecular levels. Low to moderate levels of miR156 improved drought tolerance in alfalfa by silencing SPL13, increasing accumulation of stress mitigating metabolites such as proline, GABA, anthocyanins and other flavonoids, as well as enhancing photosynthetic assimilation rate, Fv/Fm ratio and root development. Moreover, transcripts mediating secondary metabolite accumulation and photosynthesis such as MYB112, PAL, DFR, FGT2, PSI and PSII were increased significantly in moderate overexpressors of miR156. We also demonstrated that SPL13 protein binds to the promoter region of DFR to regulate its expression level. We conclude that a moderate increase in miR156 levels (0.5 to 1.5) is sufficient to enhance drought resilience in alfalfa, but higher miR156 overexpression may result in drought susceptibility. [email protected]

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AB2.5: MicroRNA guided post-transcriptional gene regulation in response to heat stress in wheat plants Sridhar RAVICHANDRAN1, Raja RAGUPATHY2, Tara EDWARDS1, Stephen ROBINSON2, Michael DOMARATZKI3, Sylvie CLOUTIER1 1. Ottawa Research and Development Centre, Agriculture and Agri-Food Canada, Ottawa, ON 2. Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, Saskatchewan, SK 3. Department of Computer Science, University of Manitoba, Winnipeg, MB MicroRNAs (miRNAs) are small (21-24bp), non-coding, regulatory RNAs that direct gene expression at the post-transcriptional level. With rising global temperature, understanding plants’ adaptation to heat stress has implications in plant breeding. In this study, the small RNA, mRNA (RNA-Seq) and degradome (parallel analysis of RNA ends) of leaf tissues harvested from control and heat stressed wheat plants were analysed. The small RNA sequences were aligned to previously annotated precursor miRNA to identify 208 miRNAs, of which 47 were differentially expressed after heat stress. RNA sequencing identified a total of 32,428 transcripts, of which 7634 were differentially regulated in response to heat stress. Clustering analysis of the differentially expressed miRNAs and mRNA transcripts revealed highly altered expression profiles immediately after heat stress that returned to levels similar to the control plants at one and four days after the end of the heat stress. Transcripts upregulated with heat stress were predicted to encode proteins associated with signal transduction pathways, antioxidant and methyltransferase activity. Through degradome sequencing, 64 of the differentially expressed transcripts were identified to be regulated by miRNA-mediated cleavage. Accurate identification and validation of miRNAs and their target genes are essential to develop novel regulatory gene-based breeding strategies. [email protected]

AB2.6: Association mapping of genes conferring high winter survival in autumn-seeded rye (Secale cereale L.) Hirbod Bahrani1*, Monica Baga1, Jamie Larsen2, Ravindra N. Chibbar1 1. Department of Plant Science, University of Saskatchewan, Saskatoon, SK, S7N 5A8, Canada 2. Lethbridge Research and Development Centre, Agriculture and Agri-Food Canada, Lethbridge, AB, T1J 4B1, Canada

Plants undergo cold acclimation during the fall season in preparation for freezing temperatures during winter. This process allows the most hardy rye cultivars to acquire very high resistance to frost (LT50 = -33°C) as compared to winter wheat (LT50 = -23°C). Thus, the winter rye survival on the Canadian Prairies is relatively good in contrast to winter wheat. There have been many unsuccessful attempts to transfer the winter hardiness character of winter rye to wheat through breeding. To improve frost resistance in wheat, a complete understanding of the frost resistance regulon in rye is needed. This information will be useful for development of winter wheat cultivars with enhanced cold tolerance for the Canadian Prairies. In this study, a set of 96 rye genotypes with very high to low winter hardiness were selected to identify genomic regions / genes that influence cold tolerance in winter rye. Field trials with the genotypes were conducted at three locations with relatively mild, harsh, and very harsh winters to determine the plant’s field survival index (FSI). Selected developmental traits associated with frost resistance were also studied. DNA extracted from the 96 genotypes was genotyped by sequencing to identify single nucleotide polymorphisms (SNP), assembly of rye genetic maps and QTL mapping. Association mapping studies revealed that rye lines could be divided in to four subpopulations. Several marker-trait associations were identified that need to be further confirmed by ongoing field trials.

[email protected]

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AB2.7: Nitrogen source and availability alter the effect of low CO2 on plant growth and N dynamics André G DUARTE1*, Fred J LONGSTAFFE2, Danielle A WAY1,3 1. Department of Biology, The University of Western Ontario, London, ON, Canada 2. Department of Earth Sciences, The University of Western Ontario, London, ON, Canada 3. Nicholas School of the Environment, Duke University, Durham, NC, USA Low atmospheric CO2 concentrations dominated the recent evolutionary history of plants. However, our knowledge of plant performance in low CO2 environments is poor, and that of how low CO2 interacts with nitrogen source and availability is even poorer. Here, we investigated growth, photosynthetic traits and nitrogen isotopic signature in Elymus canadensis, grown from seed at either current CO2 concentrations (400 ppm) or values representative of the last glacial maximum (180-190 ppm), in combination with either high or low nitrogen supply; with the high nitrogen treatment supplied as nitrate, ammonium, or a mix of both. Low CO2 reduced total biomass by almost half, except when nitrogen was limited, where growth was unaffected by CO2. Plants grown with nitrate, ammonium, or at low nitrogen up-regulated photosynthesis at low CO2, presenting higher maximum electron transport rates (Jmax), but not maximum Rubisco carboxylation rates (Vcmax), than plants grown at ambient CO2. Growth at low CO2 also increased leaf nitrogen concentrations. Compared to plants at ambient CO2, low-CO2 plants were enriched in 15N (1.2‰) under nitrate fertilization, but 0.8‰ depleted under low nitrogen availability. Our work demonstrates that the effects of low CO2 on plants varies depending on nitrogen source and availability; such responses should be considered when inferring data about plant responses to past climates, especially when carbon resources were most limiting. André G Duarte – [email protected]

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Biotic Stress Wednesday July 5 3:20-4:50pm; FSC Room 1001 Chair: Xin Li, University of British Columbia

BS.1: Verticillium dahliae disease resistance and the regulatory pathway for tuberization in potato

Helen H. Tai1*, David De Koeyer1, Mads Sønderkær2, Sanne Hedegaard2, Martin Lägue1, Claudia Goyer1, Lana Nolan1, Charlotte Davidson1, Kyle Gardner1, Jonathan Neilson1, Jamuna Paudel1, Agnes Murphy1, Benoit Bizimungu1, Hui Ying Wang3, Xingyao Xiong3, Dennis Halterman3, and Kåre Lehmann Nielsen2 1Agriculture and Agri-Food Canada, Fredericton Research and Development Centre, P.O. Box 20280, Fredericton, New Brunswick, Canada E3B 4Z7 2 Department of Chemistry and Bioscience, Fredrik Bajers Vej 7, Aalborg University, 9220 Aalborg, Denmark 3 College of Horticulture and Landscape, Hunan Agriculture University, Hunan, Changsha, China 410128 Verticillium dahliae Kleb. is a pathogenic fungus causing wilting, chlorosis and early dying in potato. Genetic mapping of resistance V. dahliae was done using a diploid population in potato. A quantitative trait locus (QTL) on chromosome 9 was identified that included the Verticillium wilt resistance gene was mapped using the Ve2 marker and was found to be located within the chromosome 9 QTL. The major QTL was found on chromosome 5. The StCDF1 gene controlling maturity and tuberization was mapped within the interval. Epistasis analysis indicated that the two loci on chromosomes 5 and 9 had highly significant interaction, and that StCDF1 functioned downstream of Ve2. Expression QTL (eQTL) analysis was carried out. Gene Ontology (GO) analysis was done and genes with eQTL at the StCDF1 and Ve2 loci were both were found to have similar functions involving the chloroplast including photosynthesis. However, differences were also noted. Among the GO terms that were specific to genes with eQTL at the Ve2, but not the StCDF1, locus, were those associated with fungal defense. The expression of genes regulated by StCDF1 and functioning downstream in the pathway for regulation of maturity and tuberization was also genetically mapped. The mobile tuberigen, StSP6A, and the gene upstream of it, StSP5G, were found to have an eQTL on chromosome 5 at the same location as that found for V. dahliae resistance. Together these results suggest that the gene pathway controlling maturity and tuberization is involved in V. dahliae infection. Helen Tai [email protected] mail.ca BS.2: Negative Regulation of Plant Immunity by the ANP2/ANP3-MKK6-MPK4 kinases cascade

Rowan van Wersch*, Kehui Lian, Fang Gao and Yuelin Zhang Department of Botany, University of British Columbia, Vancouver, BC, Canada Arabidopsis MPK4 is a component of two independent MAP kinase cascades and functions in regulating development as well as plant defense. The MEKK1-MKK1/MKK2-MPK4 cascade inhibits the activation of defense responses mediated by the NB-LRR protein SUMM2, whereas the ANPs-MKK6-MPK4 cascade plays an essential role in cytokinesis. Here we report a novel role for the ANPs-MKK6-MPK4 cascade in regulating plant immune responses. Loss of function of MKK6 or ANP2/ANP3 results in constitutive activation of plant defense responses. The autoimmune phenotypes in mkk6 and anp2 anp3 mutant plants can be largely suppressed by a constitutively active mpk4 mutant. Further analysis showed that constitutive defense response in anp2 anp3 is dependent upon the defense regulators PAD4 and EDS1, but not SUMM2, suggesting that the ANP2/ANP3-MKK6-MPK4 cascade negatively regulates a SUMM2-independent defense response pathway. To better understand how the ANP2/ANP3-MKK6-MPK4 cascade regulates immune responses, we carried out a suppressor screen in the anp2 anp3 double mutant background. A large number of mutants that suppresses the dwarf morphology and constitutive immune responses in anp2 anp3 were identified. Analysis of these suppressor mutants of anp2 anp3 will be presented. Rowan van Wersch [email protected]

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BS.3: A novel finding links the Ca2+ channel activity of CYCLIC NUCELTOIDE GATED CHANNEL 2 to auxin signaling

Sonhita CHAKRABORTY1*, Masatsugu TOYOTA2,3, Wolfgang MOEDER1, Simon GILROY3, Keiko YOSHIOKA1*

1. Department of Cells and Systems Biology, University of Toronto, Toronto, ON, Canada 2. Graduate School of Science and Engineering, Saitama University, Saitama City, Saitama, Japan 3. Department of Botany, University of Wisconsin, Madison, WI, USA

Cyclic Nucleotide Gated Channels (CNGCs) are non-selective cation channels that are involved in regulating abiotic and biotic stress responses in plants. defense, no death1 (dnd1), the CNGC2 null mutant, exhibits autoimmune phenotypes and Ca2+ hyper-sensitivity, indicating that CNGC2 is a Ca2+ conducting channel that is involved in plant immunity. In order to understand CNGC-mediated signaling, the first CNGC2 suppressor, repressor of defense, no death1 (rdd1), was identified. Through a combination of conventional map-based cloning and whole genome sequencing, the rdd1 mutation was unexpectedly found to be a point mutation in an auxin biosynthesis gene. This discovery led us to re-phenotype dnd1 from auxin point of view. Interestingly, not only does dnd1 exhibit lower levels of endogenous auxin in the root, it also displays partial insensitivity to exogenous auxin. These alterations in auxin signaling in dnd1 suggest that CNGC2 is likely involved in auxin-related processes. Furthermore, using the fluorescence resonance energy transfer (FRET) based genetically encoded Ca2+ indicator YC-nano65, we demonstrate for the first time that dnd1 plants exhibit an impaired Ca2+ influx after auxin treatment. Taken together, our data indicate that beyond its known function in plant immunity CNGC2 is also involved in auxin signaling through its Ca2+ channel activity. More data connecting the CNGC2 to auxin signaling will be presented.

Sonhita Chakraborty - [email protected] BS.4: Elucidating the immune functions of plant TRAF domain proteins using CRISPR Kevin Ao1,2, Shuai Huang1,2,3, Jianhua Huang1,2, Xin li1,2 1. Michael Smith Laboratories, University of British Columbia, Vancouver, BC 2. Department of Botany, University of British Columbia, Vancouver, BC 3. Howard Hughes Medical Institute/Yale University School of Medicine, Yale University, New Haven, CT Plants encounter and defend against a wide variety of pathogens daily. Due to a robust immune system, most plants thrive despite the constant biotic challenges they are pit against. However, successful plant pathogens have evolved mechanisms to overcome the first layer of plant immune receptors. Intracellular receptors that recognize these mechanisms form a second layer of defense that provide plants an advantage. Although much progress has been made in our lab regarding the post-transcriptional/translational modification, the transport, or the degradation of these intracellular receptors, it is still unclear how downstream signaling pathways are activated. One proposed reason for why the missing regulatory genes have not been found through forward genetic screens, is that these regulators have multiple copies and are redundant. One family of genes containing TRAF (TNF Receptor Associated Factors) domains, have members that exists in tandem repeats and duplications. The TRAF gene family is one appealing candidate since human equivalents are involved in innate immunity signaling pathways, and since one pair of redundant TRAF genes was recently found to be involved in Arabidopsis immunity. With the development of CRISPR-CAS9 as a genome-editing tool, we carried out a reverse genetic screen to create knock-out mutations in multiple clades of this large family of genes. Here we present two candidates and preliminary characterization of their immune phenotypes. Further studies in this area, will improve our understanding of the plant immune system and advance goals to breed superior crops against the rise of virulent plant pathogens. Kevin Ao [email protected]

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BS.5: Identification and functional characterization of putative effectors of Plasmodiophora brassicae and their role in regulating cell death during infection. Md. Musharaf HOSSAIN1*, Yangdou WEI1 & Peta C. BONHAM-SMITH1 Department of Biology, University of Saskatchewan, Saskatoon, SK. Plasmodiophora brassicae is a devastating obligate biotrophic pathogen that causes clubroot disease in Brassicas. Once inside a host cell, the pathogen secretes effector proteins to manipulate cellular processes and facilitate colonization. In this study, the expression of 75 putative secretory effector protein coding genes were studied at different time points; 0, 2, 5, 7, 14, 21 and 28 days post infection (dpi) of A. thaliana Col-0 with P. brassicae pathotype 3. Fourteen of the 75 genes were not expressed in resting spores of P. brassicae but were expressed at later stages of infection/disease progression, suggesting that these genes may be associated with pathogenicity and disease progression. Many of the 75 genes showed significant up- or down-expression over the time course of this study. The distinctive expression patterns, specific to certain infection stages, of some of the 75 genes suggest an importance during primary and secondary infection by P. brassicae. To date, 46 of the genes have been screened for their ability to inhibit PiNPP1-induced programmed cell death (PCD) using an Agrobacterium-mediated co-infiltration method. Fourteen P. brassicae putative effectors have been selected, based on their expression pattern and PCD challenge result, for further study. Future work will identify the role of some of these putative effectors in minimising the plant response to the pathogen. [email protected] BS.6: Arabidopsis Transcription factors TGA1 and TGA4 regulate salicylic acid and pipecolic acid biosynthesis by modulating the expression of SARD1 and CBP60g Tongjun Sun1, Lucas Busta1,2, Qian Zhang1, Pingtao Ding1, Reinhard Jetter1,2 and Yuelin Zhang1 1. Department of Botany, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 2. Department of Chemistry, University of British Columbia, Vancouver, BC V6T 1Z4, Canada The Arabidopsis transcription factors TGA1 and TGA4 are involved in basal resistance against pathogens, but the mechanism of how they regulate plant immunity is unknown. On the other hand, salicylic acid (SA) and pipecolic acid (Pip) are two plant metabolites with important roles in defense signaling, and their levels increase dramatically following pathogen infection. In this study, we tested whether TGA1 and TGA4 regulate pathogen-induced SA and Pip biosynthesis. First, we found that both TGA1 and TGA4 modulate the expression of SARD1 and CBP60g, which encode two master transcriptional regulators of plant immunity, thus promoting pathogen-induced SA biosynthesis. Second, we found that SARD1 and CBP60g also positively regulate Pip biosynthesis by targeting genes encoding key enzymes involved in the process. In plants overexpressing SARD1, genes encoding Pip biosynthesis enzymes are up-regulated, and Pip abundance is dramatically increased. Furthermore, pathogen-induced Pip accumulation is greatly reduced in the sard1 cbp60g double mutant. In tga1 tga4 mutant plants, the expression levels of SARD1 and CBP60g along with SA and Pip accumulation following pathogen infection are significantly reduced compared to wild type plants. Overall, our data strongly suggest that TGA1 and TGA4 positively regulate plant immunity by promoting pathogen-induced SARD1 and CBP60g expression. Tongjun Sun [email protected]

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Cell Biology II (Cell Walls) Thursday July 6 1:30-3:00pm; FSC Room 1005 Chair: Miki Fujita, University of British Columbia CB2.1: The biomechanical role of cellulose-enriched secondary cell walls in a wave-swept seaweed Patrick T. MARTONE1, Kyra JANOT1, Miki FUJITA1, Katia RUEL2, Jean-Paul JOSELEAU2, Hwan Su YOON3, Debashish BHATTACHARYA4, José M. ESTEVEZ5 1. Department of Botany and Biodiversity Research Centre, University of British Columbia, Vancouver, BC, V6T 1Z4 CANADA 2. E.I. LINK-Conseil, 349 rue du Mont-Blanc, 38570 Le Cheylas, France 3. Department of Biological Sciences, Sungkyunkwan University, Suwon 16419, Republic of Korea 4. Department of Ecology, Evolution and Natural Resources, Rutgers University, New Brunswick, New Jersey 08901, USA 5. Fundación Instituto Leloir and Instituto de Investigaciones Bioquímicas Buenos Aires, Consejo Nacional de Investigaciones Científicas y Técnicas (IIBBA-CONICET), C1405BWE, Buenos Aires, Argentina Cellulose-enriched secondary cell walls are the hallmark of woody vascular plants, which develop thickened walls to support upright growth in air and resist toppling. Cellulosic secondary walls evolved independently in the red seaweed Calliarthron, presumably to reinforce thalli against forces applied by crashing waves, but the biomechanical role of these secondary walls is poorly known. Calliarthron possesses two CesA gene sequences that are phylogenetically distinct from those in land plants and, like CesA’s from other red algae, contain a carbohydrate-binding module (CBM48) of unknown function. Secondary walls in Calliarthron contain half as much cellulose (~22% w/w) as secondary walls of land plants (e.g. Arabidopsis), although primary walls of these taxa are similar (~8% w/w). Secondary walls in Calliarthron are similar in thickness to primary walls (~0.6 m), unlike secondary walls in vascular plants, which can be significantly thicker (up to 10 m). His-tagged recombinant CBM’s differentiated crystalline and amorphous cellulose in planta, revealing elevated levels of crystalline cellulose in secondary walls. Yet field emission scanning electron microscopy (FESEM) revealed that cellulose bundles are disorganized in both primary and secondary walls with no predominant orientation. Mature tissues with thick secondary walls were 20% stronger, 50% stiffer, and 60% tougher than immature tissues. However, the deposition of secondary cell walls did not impact the extensibility of tissues, which double in length before breaking and are unique among seaweeds and land plants. Chemical and structural elaboration of secondary walls likely contributes to the survival of Calliarthron populations along wave-battered coastlines throughout the NE Pacific. [email protected] CB2.2: Developing a System to Engineer Arabidopsis thaliana Seed Coat Mucilage Composition Robert MCGEE1, George HAUGHN1 1. Department of Botany, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada. The seed coat epidermal cells of Arabidopsis thaliana synthesize and secrete large quantities of mucilage, a specialized secondary cell wall composed of the three major primary cell wall components: cellulose, hemicelluloses and pectin. As a means to investigate cell wall structure and function, we are attempting to modify mucilage carbohydrate composition and and assess its impacts upon mucilage function and properties. As a first step, we fused three previously-identified seed coat-specific promoters to a known mucilage catabolic enzyme, MUCILAGE-MODIFIED2 (MUM2) and demonstrated their ability to complement a mum2 mutation. Therefore all three promoters have the potential to drive expression of carbohydrate active enzymes in a spatial and temporal pattern and at a level sufficient to manipulate carbohydrate composition. We are currently using these seed coat-specific promoters to drive expression of carbohydrate active enzymes not normally found in the seed coat epidermal cells. Enzymes were selected based on their respective ability to modify

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one of three mucilage components: homogalacturonan, rhamnogalacturonan I (RG-I) and cellulose. Initially, we have targeted two different YFP-tagged fungal RG-I degrading enzymes to the apoplast of the seed coat epidermal cells. Confocal fluorescence microscopy was used to confirm their localization. Consistent with the fact that RG-I is the major component of seed mucilage, seeds of the transgenic plants expressing either enzyme appear to have significantly lower levels of mucilage. We are currently exploring this hypothesis further by analyzing the transgenic seed both biochemically and cytologically. Robert McGee: [email protected] CB2.3: There’s Something About Pectin: Glandular Secretory Trichome Ultrastructure in Artemisia annua Rebecca A. SMITH1, David A BIRD1 1. Department of Biochemistry, and the US Department of Energy’s Great Lakes Bioenergy Research Center (GLBRC), the Wisconsin Energy Institute,University of Wisconsin-Madison, Madison, United States of America 2. Department of Biology, Mount Royal University, Calgary, Alberta, Canada Artemisia annua produces an array of isoprenoids, which are sequestered within the glandular secretory trichomes found in this medicinal plant. The glandular secretory trichomes (GSTs) have a distinct ten-celled shape and developmental pattern. During maturity, the cuticle separates from the two upper cell pair to create a subcuticular space in which the natural products accumulate. Although detached from the primary cell wall, the cuticle continues to expand in size, enlarging the subcuticular space. The phenomenon of cuticular separation to yield a subcuticular space leads to questions about the structure and composition of the GST cuticle and cell walls, particularly of the apical and second tier cells where the cuticle separates from the cell wall. Does the cuticle ‘stretch’ to accommodate the expansion of the subcuticular space? Is there a change in the structure or composition that allows the cuticle to detach from the cell wall in such a localized manner? Does the structure of the cell wall change to prevent the diffusion of natural products from the storage location back into the GST cells? In the hopes of addressing some of these questions, we undertook an in-depth microscopic examination of the ultrastructure of the GSTs in Artemisia annua. We found that, although the cuticle must expand by over 2 fold, remarkably, the overall thickness of the cuticle remains unchanged. Furthermore, using monoclonal antibodies, we identify rhamnogalactan-I epitopes specific to GST cell walls, the abundance of which change during development. How these findings address the posed questions will be discussed. David Bird, [email protected]

CB2.4: Cellulose biosynthesis inhibitors affect more than cellulose!

Fatima Awwad1*, Guillaume Bertrand2, Miki Fujita3, Michel Grandbois2, Geoff Wasteneys3, Nathalie Beaudoin1. 1. Centre Sève, Université de Sherbrooke, Sherbrooke, QC, CA 2. Institut de Pharmacologie de Sherbrooke, Université de Sherbrooke, Sherbrooke, QC, CA 3.Botany Department, University of British Columbia, Vancouver, BC, CA

Cellulose biosynthesis inhibitors (CBI) are generally synthetic compounds which compromise cell wall stability in plant cells. In contrast, thaxtomin A is a natural CBI that is synthesized by the plant pathogen Streptomyces scabies which causes common scab on potato tubers. Treatment of cell suspensions with thaxomin A induces changes in gene expression and an atypical programmed cell death (PCD) that are similar to those induced by another well-known CBI, isoxaben. However, the specific mode of action and target of thaxtomin A are still unknown. We have first investigated the mechanisms involved in the induction of PCD by CBIs. Our results show the implication of auxin transport and modulation of oxidative stress in the induction of PCD. In order to understand how thaxtomin A perturbs cellulose synthesis and cell wall organization, we have characterized its effect on cell wall stiffness using atomic force microscopy. Studies conducted on microtubule dynamics and cellulose synthase velocity in the presence of thaxtomin A have also provided new information on the mechanism of action of this natural CBI. These studies bring a deeper understanding of the specific mode of action of thaxtomin A and provide new information that may be useful for the development of new strategies in the control of common scab. Fatima Awwad [email protected]

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CB2.5: CELLULOSE SYNTHASES in Primary to Secondary Cell Wall Transition During Tracheary Xylem Development

Yoichiro Watanabe1,2*, René Schneider3, Sarah Barkwill2, Eliana Gonzales-Vigil2, Joseph Hill4, Staffan Persson3, Shawn D. Mansfield2, A. Lacey Samuels1.

1. Department of Botany, University of British Columbia, Vancouver, BC 2. Department of Wood Science, University of British Columbia, Vancouver, BC 3. School of BioSciences, University of Melbourne, Parkville, VC, Australia. 4. Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, PA.

Cellulose is the most abundant biopolymer in nature, and is synthesized by CELLULOSE SYNTHASE (CESA) Complexes (CSC) at the plasma membrane, where it extrudes cellulose into the cell wall. Cellulose quantity and quality differ significantly between primary and secondary cell walls. This variation is partially explained by the presence of multiple CESA isoforms, including unique complexes of CESAs for production of the two walls. Previous work has shown that the different CESAs are largely non-redundant with few interactions between primary and secondary cell wall CESAs. Given this, it is unclear whether the primary wall CESAs contribute to cellulose synthesis during the transition to secondary cell wall production. To clarify this, we characterized the cellular dynamics and localization of primary and secondary cell wall CESAs at the onset of secondary cell wall synthesis using fluorescently tagged CESA6 and CESA7 in developing Arabidopsis thaliana tracheary elements. Using live-cell imaging, a brief transition period was observed where both primary and secondary wall CESAs coexisted in banded plasma membrane domains, but with distinct dynamic behaviors. As secondary wall deposition increased, the majority of primary wall-specific CESAs were no longer localized on the plasma membrane, and appeared in endosomes and later lytic vacuoles. Western blotting indicated a corresponding decrease in primary CESA protein levels. Conversely, secondary wall-specific CESAs were abundantly present at the plasma membrane and Golgi. These changes reflect distinct and dynamic subcellular trafficking patterns for primary and secondary cell wall CESAs to affect the transition between primary and secondary cell wall synthesis.

Yoichiro Watanabe [email protected]

CB2.6: Mechanics of complex shape formation in plant cells Amir J. BIDHENDI1, Bara ALTARTOURI1, Anja GEITMANN2 1. Institut de recherche en biologie végétale, University of Montreal, 4101 Rue Sherbrooke Est, Montreal, Québec H1X 2B2, Canada 2. Department of Plant Science, McGill University, Macdonald Campus, 21111 Lakeshore, Ste-Anne-de-Bellevue, Québec H9X 3V9, Canada The driving force of plant cell growth is provided by the intracellular turgor pressure. However, since the turgor pressure is a scalar entity, anisotropic growth of plant cells is maintained by dynamic regulation of the mechanical properties at the cell wall level. It is known that anisotropic and/or non-uniform growth of plant cells results in a wide variety of different shapes that are intimately related to their specific function, but the mechanics underlying formation of complex shapes in plant cells is poorly understood. In eudicotyledons, pavement cells covering the surface of plant leaves form interlocking protrusions and indents resembling a jig-saw puzzle. These cells are an ideal model to study how cell wall mechanics is regulated to generate complex morphologies. In this study, we examined the distribution and status of two major cell wall polysaccharides, pectin and cellulose, through confocal laser scanning microscopy. The imaging data were used to inform finite element models of cell wall expansion to link cell wall mechanics to cellular morphogenesis. In the in silico simulation of the generation of lobed cell shapes the spatial distribution of cell wall components was found to correlate with their hypothesized roles. Importantly, a finite element model with a positive mechanical feedback loop between the stress and stiffness in the cell wall showed that infinitesimally small variations in local wall stiffness can be triggers for cell shaping if local inhibitory mechanisms are implemented. Anja Geitmann [email protected]

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Ecophysiology Thursday July 6 1:30-3:00pm; FSC Room 1001 Chair: Prakash Venglat, University of Saskatchewan EP.1: Quantitative assessments of wax-only water barriers on Arabidopsis thaliana stems and Pinus banksiana needles Christopher BUSCHHAUS1 1. Crandall University, Moncton, NB With extreme weather events predicted to increase, the contribution and plasticity of cuticular wax (as compared to other epidermal components) as a primary barrier to environmental stressors must be quantified. Most studies demonstrating that plant wax restricts excessive water loss have analyzed leaves or fruit using a chamber apparatus that maintains a saturated state at the physiologically inner surface of the cuticle. This set-up permits measuring the same leaf area before and after a treatment. Conversely, studies of narrow cylindrical plant parts such as needles or thin stems have relied on primitive dehydration techniques that do not allow further treatments on the same plant material. Here we modify the chamber apparatus to examine the minimum water conductance of needles and narrow stems in order to allow paired comparisons, such as before and after wax removal. Results using stems from Arabidopsis thaliana and needles from Pinus banksiana indicate resistances to water movement comparable to published values for other leaves followed by a substantial decrease after solvent extraction of cuticular wax. Application of this method should allow quantitative assessments of the wax (only) contribution to the water barrier in Arabidopsis wax mutants as well as the plasticity of cuticular wax water barriers necessary for extreme environmental conditions likely to be experienced by conifer needles. Christopher Buschhaus [email protected]

EP.2: A family of Arabidopsis MYB transcription factors that control the regulation of suberin deposition Hefeng HU1*, Daniel KLEIN1, Jhadeswar MURMU1, Owen ROWLAND1 1. Department of Biology and Institute of Biochemistry, Carleton University, Ottawa Suberin is a cell wall-associated polymer consisting of glycerol, phenolics, and various chain-length fatty acids and fatty alcohols. Suberin is deposited in diverse plant tissues including root exodermis and endodermis, aerial and underground periderms, and seed coats. While suberin is produced constitutively in these specialized tissues, suberization of cell walls also occurs in these and other tissues under stresses such as wounding. Suberin plays important roles in controlling water and solute movement, especially in roots. It also helps plants to defend against various stressors, including drought, high salinity, toxic metals, insects, and microbial pathogens. Although much progress has been made in identifying genes encoding suberin biosynthetic enzymes, the molecular mechanisms governing the regulated deposition of suberin are currently unclear. We provide evidence that Arabidopsis transcription factors MYB53, MYB92, and MYB93 are important regulators of suberin during root development. We first identified these MYB proteins as positive suberin regulators in a transient assay screen using leaves of Nicotiana benthamiana. Analysis of promoter::GUS transgenic lines revealed that MYB53, MYB92, and MYB93 have overlapping gene expression patterns in root endodermis when suberin is being deposited. Loss-of-function mutants of the three transcription factors exhibited major reductions of suberin in the endodermis of young roots. We also characterized an Arabidopsis steroid-inducible overexpressing MYB53 line and found that suberin can be rapidly and ectopically induced in both roots and leaves. The identification of master regulators of suberin provides the means to generate crops that are more stress resistant via enhancement of their suberized cell walls. Hefeng Hu: [email protected]

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EP.3: Abscisic acid and ethylene are integrated in the phytoglobin (Pgb) regulation of maize somatic embryogenesis Mohamed M. MIRA1, Belay T. AYELE, Robert D. HILL, Claudio STASOLLA, Karuna KAPOOR* 1. Department of Plant Science, University of Manitoba, Winnipeg, Manitoba, Canada R3T 2N2 Suppression of Zea Mays phytoglobins (ZmPbg1.1 or ZmPgb1.2) during somatic embryogenesis induces programmed cell death (PCD) by elevating nitric oxide (NO). While ZmPgb.1.1 is expressed in many embryonic domains and its suppression results in embryo abortion, ZmPgb1.2 is expressed in the basal cells anchoring the embryos to the embryogenic tissue. Removal of these “anchor cells” by PCD allows the embryos to develop further. The effects of ZmPgb suppression on embryogenesis were abolished by exogenous applications of abscisic acid (ABA). A depletion of ABA, ascribed to a down-regulation of biosynthetic genes was observed in those embryonic domains where the respective ZmPgbs were repressed via NO modulation. Depletion in ABA content induced the transcription of ethylene biosynthetic and responsive genes, as well as the accumulation of ethylene, which influenced embryogenesis. Somatic embryo number was reduced by high ethylene levels and increased with pharmacological treatments suppressing ethylene synthesis. The ethylene inhibition of embryogenesis was linked to the production of ROS and the execution of PCD. Integration of ABA and ethylene in the ZmPgb regulation of embryogenesis is proposed in a model where NO accumulates in ZmPgb-suppressing cells, decreasing the level of ABA. Abscisic acid inhibits ethylene biosynthesis and the NO-mediated depletion of ABA relieves this inhibition causing ethylene to accumulate. Elevated ethylene levels trigger production of ROS and induce PCD. Ethylene-induced PCD in the ZmPgb1.1-suppressing line leads to embryo abortion, while PCD in the ZmPgb1.2-suppressing line results in the elimination of the anchor cells and the successful development of the embryos. Karuna Kapoor [email protected]

EP.4: Canola yields under hot and dry climates Raed ELFERJANI1, Raju SOOLANAYAKANAHALLY1, Branimir GJETVAJ1 1. Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, Saskatoon, SK Extreme heat waves and soil moisture deficit are projected to impair global food security as a result of climate change. Mimicking field-like conditions inside a greenhouse, we investigated physiological acclimations of Brassica napus L. (canola) by exposing to well-watered, drought, heat and drought+heat stress combinations. Normalized difference of vegetation index (NDVI) and efficiency of photosystem II (Fv/Fm) were reduced mainly by drought+heat rather than by single stressors. Maximum carboxylation rate (Vcmax) and rate of RuBP regeneration (Jmax) were significantly decreased under heat as well as drought+heat, and to a lesser extent by drought alone, suggesting a prominent role of heat over water deficit on impairing carbon fixation apparatus and electron transport chain. Conductance of CO2 through stomata (gs) and along the mesophyll (gm) responded differently to stressors, with gm reduced by 20% and 28% on exposure to heat and drought+heat treatments, respectively. On the other hand, gs dropped significantly when exposed to water deficit. A strong positive correlation between seed yield versus Vcmax (R2=0.63), Jmax (R2=0.64), and gm (R2=0.64) was observed. Leaf δ13C and δ15N values changed significantly under drought+heat stress and a negative correlation was found between δ15N and yield. In addition, NDVI and Fv/Fm were also positively correlated with seed yield and are good indicators of stress severity in canola prior to flowering. Our preliminary study highlights the effect of heat on canola fertility in order to maintain yield security. Raed Elferjani ([email protected])

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EP.5: Arabidopsis sucrose synthases localize to phloem, not xylem, suggesting a role in phloem loading and unloading Danyu Yao1* Eliana Gonzales-Vigil1, Lacey Samuels2, and Shawn D. Mansfield1 1. Department of Wood Science, University of British Columbia, Vancouver, British Columbia, Canada 2. Department of Botany, University of British Columbia, Vancouver, British Columbia, Canada In most plant species sucrose is the primary transport sugar derived from photosynthesis, and is crucial for plant growth and development. Sucrose is key to establishing sink strength, facilitating starch and oil biosynthesis, as well as respiration and phloem loading. Sucrose synthase (SuSy), one of two enzymes that can catalytically cleave sucrose, mediates the reversible conversion of sucrose and UDP into UDP-glucose and fructose. In sink tissues such as secondary xylem, SuSy has been proposed to be involved in the synthesis of cell wall polymers, including cellulose and callose. The widely-cited model is that a plasma-membrane-located SuSy tightly associates with the cellulose synthase complex to channel UDP-glucose derived from sucrose directly to cellulose biosynthesis. In Arabidopsis thaliana, SuSy exists as six isozymes that exhibit tissue-specific expression profiles. To investigate the spatiotemporal localization of SuSy and its potential role in cellulose synthesis, Yellow Fluorescent Protein -fusions of all six SuSy were transformed into wild-type and sus1/sus4 mutant plants. Confocal microscopy of mature stems and leaves revealed phloem-cell-specific localization of all SuSy isozymes. Live-cell imaging clearly indicated that SuSy1 and SuSy4 were only confined to companion cells, while SuSy5 and SuSy6 specifically localized to phloem sieve elements. In contrast, SuSy2 was absent in the roots, stems, and leaves but was highly expressed in seeds during early maturation. SuSy3 was highly expressed in the embryo at the late maturation phase, as well as in guard cells of leaf stomata. These findings bring into question the current model of cellulose biosynthesis in Arabidopsis. Danyu Yao [email protected] EP.6: Plant meristems function as sites of integration of developmental and environmental stressors Prakash VENGLAT*, Perumal VIJAYAN, Karen TANINO Department of Plant Sciences, College of Agriculture and Bioresources, Univ. of Saskatchewan, Saskatoon, Canada Self-organizing groups of cells called meristems are initiated during embryo development and post-embryonically give rise to various tissue systems of the adult flowering plant. The embryonic poles defined by shoot and root apical meristems (SAM and RAM) give rise to the entire shoot and root systems respectively. Procambium, initiated as cellular strands between the embryonic SAM and the RAM form the vascular conducting tissue during seedling growth. The procambium transitions into a third bifacial meristem, called the vascular cambium that enables secondary growth. Branching of the shoot and the root systems are accomplished by the shoot axillary meristems and the lateral root meristems that emerge from the pericycle, itself a product of the RAM. The timing, location and rate of meristematic activity basically orchestrate the varied architecture, developmental phases of a plant’s life cycle and phenomena like dormancy and flowering. There is evidence to show that the internal developmental changes as well as external environmental changes can be sources of stress. We will present an approach for developing an integrated view of plant development in the context of meristems adapting to phase changes and environmental stresses by comparing the gene regulatory networks that regulate the function of meristems. Integrating the understanding of these mechanisms will provide a deeper knowledge base for understanding the plant phenotype, adaptive response and variability. Prakash Venglat; email - [email protected]

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Technological Innovations Thursday July 6 1:30-3:00pm; FSC Room 1221 Chair: Ingo Ensminger, University of Toronto, Missassauga TI.1: Photosynthetic phenology reflected by leaf optical properties, chlorophyll fluorescence and dynamics of the photoprotective xanthophyll cycle Ingo ENSMINGER1, Emmanuelle FRÉCHETTE1, Laura JUNKER1,2, Chris WONG1, Christine CHANG1,3 1. Department of Biology, Graduate Programs in Cell & Systems Biology and Ecology & Evolutionary Biology, University of Toronto, 3359 Mississauga Road, Mississauga, ON, Canada 2. Present address: Institute of Bio and Geosciences IBG-2, Plant Sciences, Forschungszentrum Jülich, Jülich, Germany 3. Present address: Cornell University, Department of Crop and Soil Sciences, Ithaca, NY The ability of plants to sequester carbon is highly variable over the course of the year and reflects seasonal variation in photosynthetic efficiency. This seasonal variation is most prominent during autumn, when leaves of deciduous tree species undergo senescence, which is associated with the downregulation of photosynthesis and a change of leaf color and leaf optical properties. Vegetation indices derived from remote sensing of leaf optical properties using e.g. spectral reflectance measurements are increasingly used to monitor and predict growing season length and seasonal variation in carbon sequestration. Here we show that some of the widely used vegetation indices such as the normalized difference vegetation index (NDVI) and photochemical reflectance index (PRI) vary in their ability to adequately track the seasonal variation in photosynthetic efficiency and chlorophyll content. We further show that monitoring seasonal variation of photosynthesis using NDVI or PRI is particularly challenging in evergreen conifers, due to little seasonal variation in foliage. However, there is remarkable seasonal variation in leaf optical properties associated with changes in pools of xanthophyll cycle pigments and carotenoids that provide a promising way of monitoring photosynthetic phenology in evergreen conifers via leaf reflectance measurements. Ingo Ensminger [email protected]

TI.2: Exploring the innerSPACe of plants: Insights into plant water uptake and transport from synchrotron-based X-ray microCT Andrew J. McElrone USDA-Agricultural Research Service; Department of Viticulture and Enology, University of California, Davis USA Water scarcity threatens agricultural productivity in dry growing regions throughout the world. A primary cause of lost plant productivity under drought is failure to absorb and transport adequate amounts of water to a transpiring canopy, however, the exact site of this failure is still being debated. We have utilized synchrotron-based X-ray micro-computed tomography (microCT), a non-destructive imaging technique, to provide new insights into water uptake and transport capacity in living plants subjected to drought stress. I will provide a synopsis of our work from the last decade. Our original efforts were aimed at reconstructing and modelling network properties of the xylem to better understand how embolism (i.e. air bubble blockages) and vascular pathogens spread systemically in plants. The 3D nature of the datasets enabled the discovery of unique anatomical structures that help to explain unusual network properties and differences among genotypes that vary in susceptibility to stress. Our previous in vivo studies, which describe patterns of drought-induced embolism formation, spread, and repair across species, will be compared with new datasets revealing weak links in the hydraulic system and dynamic changes in tissue water content. I will discuss how results differ if measurements were done on intact or excised plant samples, and implications for the interpretation of plant and organ total water use and how various compartments contribute to an integrated response to plant stress. [email protected]

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TI.3: Synchrotron based imaging: Emerging technologies for agricultural research and innovation Jarvis A. STOBBS1, Chithra KARUNAKARAN2, Rachid LAHLALI3, Toby BOND4, Dean CHAPMAN5 1. Environmental and Earth Sciences - Science Division, Canadian Light Source, Saskatoon, SK 2. Environmental and Earth Sciences - Science Division, Canadian Light Source, Saskatoon, SK 3. Agronomy, Plant Protection and Animals Health Environmental Sciences - Ecole Nationale d'Agriculture de Meknès, Meknès, Morocco 4. Industrial Science – Canadian Light Source, Saskatoon, SK 5. Science Director - Science Division, Canadian Light Source, Saskatoon, SK The Canadian Light Source (CLS) is a national research facility, one of the largest science projects in our country’s history, and the brightest light in Canada—millions of times brighter than even the sun—used by over 1,000 scientists from around the world every year in ground-breaking health, environmental, materials, and agricultural research. One of the unique strengths of synchrotron techniques is the study of internal structures of live plants and soil-root systems using high resolution and fast X-ray imaging methods. This enables longitudinal and dynamic studies such as uptake and movement of nutrients, and root development in plants. Synchrotron based mid infrared, soft X-ray, and hard X-ray spectromicroscopy techniques enable the study on the presence and distribution of biomolecules (lipids, proteins, polysaccharides) and nutrients (boron, nitrogen, calcium, potassium, phosphorus, sulphur, manganese, iron, zinc etc.) in plant parts such as roots, stem, leaves, seeds, and in hydrated soils. The CLS is one of the leading synchrotrons in the world that has developed and optimized synchrotron techniques for plant and soil sciences research. Further, CLS has unique suite of experimental stations ideal for the characterization of plant or soil samples from macro (millimeter scale) through cellular (micron scale) to sub-cellular (nano scale) level. Integration of structural and biomolecular imaging of plants is a novel and rapid method of identification of targeted traits for specific crop performance. Examples will be shown on how this emerging technology is being used to understand the mechanism of superior performing genotypes related to abiotic and biotic stresses in plants. Jarvis Stobbs [email protected]

TI.4: The influence of light spectral quality on basil, strawberry, and cannabis secondary metabolism. David HAWLEY*1,2, Mike STASIAK1, Mike DIXON1 1. School of Environmental Science, University of Guelph, Ontario Controlled environment agriculture strives to produce the best quality of plants as efficiently as possible. In the context of LED lighting systems in controlled environments, the assumption often is that whatever light quality yields the most biomass is the optimal system to use. Research at the University of Guelph aims to expand on this assumption by taking light spectra known for biomass production in basil, and studying the influences of these spectra on basil, strawberry, and cannabis metabolomes. Emphasis is placed on metabolites relevant to flavor, nutrition, and medicine. Analysis of metabolome allows for further optimization of spectral quality, such that the “best” light spectra for a given species can be utilized in controlled environment agriculture. Data on basil, strawberry, and cannabis metabolomes will be presented. Dave Hawley [email protected]

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TI.5: Proximity-dependent biotin identification (BioID): a novel tool for plant proteomics Madiha Khan1, 2, Rajagopal Subramaniam1, 2, Darrell Desveaux1, 3

1. Department of Cell and Systems Biology, University of Toronto, Toronto, ON, Canada M5S 3B2 2. Agriculture and Agri-Food Canada, Ottawa, ON, Canada K1A 0C6 3. Centre for the Analysis of Genome Function and Evolution, University of Toronto, ON, Canada M5S 3B2 Proximity-dependent biotin identification (BioID) has emerged as a powerful tool for studies of proteome architecture, including insoluble or membrane-associated proteins. BioID uses a promiscuous biotin ligase BirA*, translationally fused to a bait of interest to map proximally located proteins. The technique has been well established in mammalian cells, but has yet to be applied to plant systems. The application of proteomic approaches in plants poses unique challenges relative to animal systems, including low cytoplasmic volume relative to cell wall mass, high protease and phosphatase content and the predominance of ribulose-1,5-biphosphate carboxylase/oxygenase (rubisco) which can interfere with protein detection and identification. Here we present the application of the BioID technique to the model plant Arabidopsis thaliana to identify biologically relevant targets of virulence proteins from the phytopathogen Pseudomonas syringae. Madiha Khan - [email protected] TI.6: The challenge of measuring calcium uptake and distribution and its relationship with bitter pit in apple (Malus domestica Borkh) Lee A. KALCSITS1*, Luca GIORDANI1, Gregory VAN DER HEIJDEN2, David MCNEAR3 1Washington State University, Department of Horticulture, Tree Fruit Research and Extension Center, 1100 North Western Ave., Wenatchee, WA, USA 2INRA de Nancy, Rue d'Amance, 54280 Champenoux, France 3University of Kentucky, Department of Plant and Soil Sciences, Agricultural Sciences Building, Lexington, KY, USA Mineral balance is a key factor that affects quality and storability and is especially important for fleshy horticultural crops such as apple, pear, tomato, pepper and potato. Many agricultural crops are especially susceptible to physiological disorders that stem from a low concentration of calcium in the tissue, which reduces cell wall strength, resistance to biotic and abiotic stress, and necessary cell signalling. Bitter pit in apple is among these calcium -related disorders and renders 5-10% of harvested fruit unmarketable each year. For more susceptible cultivars such as ‘Honeycrisp’, it is not uncommon to lose 30-40% of the fruit to this disorder – the subsequent losses exceed $100 million in the U.S. alone. Current methods to restore calcium balance in fresh produce include the direct application of calcium to soil, the plant, and/or the fruit in addition to post-harvest treatments. However, these low-tech, top-down approaches have only provided incremental improvements and calcium disorders still plague the produce industry. To study calcium uptake and distribution in apple; a combination of novel field-based measures of elemental profiles (portable x-ray fluorescence (XRF) and calcium uptake (44Ca tracing) combined with fine-scale measurements of fruit microstructure (µ-CT) and elemental distribution (µ-XRF and µ-XANES) using synchrotron-based imaging approaches were used to shed light on the underlying mechanisms contributing to the development of bitter pit in apple in addition to other analogous problems in horticultural crops. These approaches have the potential to accelerate efforts to improve the understanding of the underlying mechanisms of calcium-related disorders in tree fruit. Lee Kalcsits [email protected]

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Specialized Metabolism II Thursday July 6 1:30-3:00pm; FSC Room 1003 Chair: Raju Soolanayakanahally, AAFC, Saskatoon SM2.1: Characterization of codeinone reductase variants in Papaver somniferum Mehran DASTMALCHI1, Peter J. FACCHINI1 1. Department of Biological Sciences, University of Calgary, AB, Canada Benzylisoquinoline alkaloids (BIAs) are a large, structurally diverse class of nitrogen-containing plant specialized metabolites. They include narcotic analgesics, codeine and morphine, found prominently in opium poppy (Papaver somniferum). Morphine biosynthesis has been largely elucidated; however, attempts at reconstituting the pathway in microbial hosts have led to low yields. A major bottleneck in the formation of morphine in engineered biosystems involves codeinone reductase (COR), which catalyzes the reversible reductions of codeinone to codeine, and morphinone to morphine. In engineered yeast, the related compounds neopine and neomorphine, not detected in the plant, are produced, diverting flux from the desired output. We have characterized five COR isoforms with high (94-97%) amino acid sequence identity. These isoforms display a remarkable, and previously undetected, activity spectrum, whereby either a majority of neopine or codeine is produced, with intermediate isoforms yielding a mixture of the two. Similarly, cell-free plant latex extract incubated with codeine or codeinone tends towards the accumulation of neopine. Outside of the plant cell context, the inherent catalytic abilities of CORs yield large amounts of undesired products that do not accumulate in the plant. Further, a substrate range experiment revealed that neopine and neomorphine could not be catalyzed by any of the CORs or plant latex extract. Therefore, these isomers represent a catalytic sink yielding a significant diversion of output over time. Site-directed mutagenesis and crystallography are being used to dissect structural variations responsible for differential production of neopine in COR variants and overcoming this bottleneck in engineered microorganisms. Mehran Dastmalchi, [email protected]

SM2.2: Virus induced gene silencing (VIGS) system for functional genomics in betalainic species, Amaranthus tricolor Dinesh ADHIKARY1*, Michael DEYHOLOS1 1. Department of Biology, The University of British Columbia, Okanagan Campus Amaranth has been cultivated for thousands of years in Central and South America. Over 70 species are known, and only three of them are domesticated. Most of the species, including Amaranthus tricolor, are rich in betalains. These red or yellow pigments are evident in different plant organs, including flower, leaf, stem, root, and cotyledon. The ecological role and nutritional and economic potential of these pigments have not been fully explored. To this end, we have silenced selected genes in the betalain biosynthetic pathway using virus induced gene silencing (VIGS). We observed that when cytochrome P450 CYP76AD1 was silenced in normally red plants, the biosynthetic intermediate L-DOPA accumulated, betacyanin pigmentation was lost in the plant, and petiole and leaf turned green. These results help to define the genetic basis of betalain synthesis in amaranths. Furthermore, this is the first report of VIGS in amaranths and demonstrates the potential of this technique for basic and applied research in these species. Dinesh Adhikary [email protected]

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SM2.3: Biosynthesis of Montbretin A: A Novel Anti-Diabetic Compound from Crocosmia Jenny(Seohyun) JO1,2*, Sandra IRMISCH1, Christopher ROACH1, Stephen G. WITHERSWithers1,3 Jörg BOHLMANN1,2 1. Michael Smith Laboratories, University of British Columbia, Vancouver, BC 2. Department of Botany, University of British Columbia, Vancouver, BC 3. Department of Chemistry, University of British Columbia, Vancouver, BC Plant metabolites have been employed by humans for centuries in traditional and modern medicine, and they remain an important source for the discovery of novel pharmaceuticals and nutraceuticals. Montbretin A (MbA), an acylated flavonoid glycoside was discovered in the underground storage organs, called corms, of the ornamental plant Crocosmia. This unique metabolite is a highly specific inhibitor of the human pancreatic α-amylase (HPA), a key enzyme in starch degradation, thus turning MbA into a promising drug candidate for the treatment of type II diabetes, a growing worldwide epidemic. However, due to its low abundance in planta, and its complex chemical structure, field cultivation and chemical synthesis, respectively, are not feasible to provide sufficient amounts of MbA to employ it as an anti-diabetic drug. Metabolic engineering of microbial or plant production systems is the most likely viable solution of the MbA supply problem. This however, requires fundamental knowledge of the biosynthetic pathway of MbA in Crocosmia. We applied a combined approach of transcriptomics, proteomics, metabolomics and biochemistry to elucidate MbA biosynthesis in the non-model plant Crocosmia. Through profiling MbA accumulation patterns throughout a growing season we could identify development specific accumulation patterns of MbA. Using this information, we were able to identify key intermediates in MbA biosynthesis along with the genes and enzymes responsible for their formation.

Jenny(Seohyun) Jo [email protected]

SM2.4: In vitro production of secondary metabolites from selected Himalayan medicinal plants Shyamal K. NANDI, Indra D. BHATT, Pitamber P. Dhyani G.B. Pant National Institute of Himalayan Environment and Sustainable Development, Kosi-Katarmal, Almora-263 643, Uttarakhand, India Himalayan medicinal plants form an integral and essential part of lives of local inhabitants who depends for their health care and livelihood. These plants are used in different system of medicine since ancient times and source of diverse array of phytochemicals which are presently being used as drugs and nutraceuticals worldwide. Due to ever increasing global demand of these compounds, the pertinent species are being subjected to reckless and destructive harvesting from the wild, often illegal, consequently many being listed under various threat categories of IUCN. In view of the above, in vitro technologies, well established for recovery of plant species, are being used for mass multiplication and subsequent plantation for cultivation, as well as culturing cells/tissues for production of secondary metabolites. The presentation will focus on selection of elite plants and using them to develop cell suspension and Agrobacterium-mediated transformed cell cultures for in vitro production of bioactive compounds in some selected herbs (Habenaria edgeworthii, Nardyostachys jatamansi, Picrorhiza kurrooa, Valeriana jatamansi, etc) of the Indian Himalayan region. In general, it was observed that the active ingredient content of in vitro cultures were found to be higher and/or comparable with that of wild populations, and hence can be used effectively for commercial applications. This will not only reduce the pressure on the existing stock but also help in the conservation of these species. The results and recent advancements made for the production of compounds of medicinal and nutraceutical value from these plant species will be highlighted.

Email: [email protected]

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M2.5: The Molecular and Biochemical Characterization of cis-Prenyltransferase Binding Protein and cis-Prenyltransferases in Parthenium argentatum with an Emphasis on Natural Rubber

Adam M Lakusta1*, Moonhyuk Kwon1, Eun-Joo G Kwon1, Dae-Kyun Ro1

1. Dept. of Biological Sciences, University of Calgary, Calgary, AB, Canada. Natural Rubber (NR) is a biopolymer with an irreplaceable role in the fields of medicine, manufacturing, and transport. Despite its necessity, the Brazilian Rubber Tree (Hevea brasiliensis) remains virtually the sole source of NR. As >90% of NR plantations are in Southeast Asia, its production is threatened by climate and disease. While NR supply remains insecure, biochemical studies have identified both a likely mechanism for NR biosynthesis and two essential proteins: cis-prenyltransferases (CPTs) and cis-prenyltransferase binding proteins (CBPs). Our research is focused on the NR production alternative guayule (Parthenium argentatum) of the Asteraceae family. CPT and CBP homologs PaCBP1 and PaCPT1-3 were identified through publically available transcriptomics databases and various biochemical assays were undertaken. Split ubiquitin and co-immunoprecipitation assays have indicated strong levels of interaction between PaCPTs and PaCBP1. Interestingly, our findings also show that guayule CPTs, together with CBP, are responsible for the production of, not NR, but a dolichol-like (C55-110) polymer in vitro. One way this was demonstrated was through the ability of CPT and CBP, only when co-expressed, to rescue the lethal phenotype of dolichol synthase-deficient yeast. Strikingly, phylogenetic and expression analyses of PaCPTs revealed PaCPT2 as both likely involved in NR biosynthesis and highly expressed throughout the plant. PaCPT2 may therefore prove to be a suitable target for future NR studies and biotechnological applications. Taken together, our data provide further evidence of a NR biosynthetic mechanism conserved across Asteraceae, while also increasing guayule’s viability as both a production alternative and model for the study of NR.

Adam Michael Lakusta [email protected]

SM2.6: Seabuckthorn: insights into the new superberry Parul JAIN1, Raju SOOLANAYAKANAHALLY1, 2, Isobel PARKIN1 and William SCHROEDER2 1. Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, Saskatoon, SK 2. Indian Head Research Farm, Agriculture and Agri-Food Canada, Indian Head, SK Seabuckthorn (Hippophae spp.) is a golden bush with unique medicinal, nutraceutical and ecological properties. Originating in Eurasia this berry bearing shrub was introduced to the Canadian prairies in the early 1930s to prevent soil erosion, improve soil health via N-fixation and enhance wildlife habitat in shelterbelts. At present, Agriculture and Agri-Food Canada has the largest in-situ seabuckthorn germplasm collection in North America representing accessions sourced from around the world. To unlock its genetic diversity and accelerate breeding, we have assembled a de novo transcriptome of developing berry and developed KASP SNP markers to differentiate male and female plants. Simultaneously, efforts are being made to select and release new cultivars with improved quality attributes. The seeds of seabuckthorn berries are rich in linoleic (~37%) and linolenic acid (~32%), while pulp oil contains a high level of palmitic (~35%) and palmitoleic acid (~40%). Overall, seabuckthorn represents a new diversification opportunity for Canadian farmers as a value added crop on marginal lands. Raju Soolanayakanahally [email protected]

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Development Thursday July 6 3:20-4:50pm; FSC Room 1221 Chair: Jae-Hyeok Lee, University of British Columbia DV.1: Complexity of the regulatory networks for the homeobox-dependent zygospore program during the sexual development of Chlamydomonas reinhardtii Sunjoo JOO, Thamali Kariyawasam, Ming Hsiu Wang, and Jae-Hyeok Lee* Department of Botany, University of British Columbia, Vancouver, BC Sexual reproduction, one of the key inventions shared by all major eukaryotic lineages, entails the cycling of two programs: gametic cell fusion generating diploids, and meiosis producing haploid progeny. C. reinhardtii has been studied as a unicellular model for this process, documenting molecular details of sex-specification, gamete-recognition/fusion, and the control of zygospore differentiation. To identify the regulatory networks orchestrating the whole sexual development, we use multiple approaches including transcriptome analysis, mutagenesis screen, and candidate mutant characterization. Zygote-specific transcriptome has revealed the molecular signature of the zygospore wall and major shifts in the flow of nucleotide-sugars and the secretory system. The pool of zygote-defective mutants provides critical tools to discover the hierarchical nature of the zygospore developmental program. Lastly, the reverse genetics approach allows interrogation of the well-known developmental regulators such as small RNA-dependent mechanisms and polycomb repressor complexes during the sexual development. Emerging complexity of the regulatory networks for the sexual development of Chlamydomonas provides plausible evolutionary paths for the complex multicellularity requiring a large number of developmental mechanisms. Jae-Hyeok Lee: [email protected]

DV.2: Cellular distribution and developmental patterns of the Crucifer ‘glucosinolate-myrosinase’ defense system Meng Li1,2*, Christoph Crocoll2, Barbara Ann Halkier2, Fred D Sack1, Carl J Douglas1, Geoffrey O Wasteneys1

1. Department of Botany, University of British Columbia, British Columbia, Canada 2. DynaMo Center, Department of Plant and Environmental Sciences, University of Copenhagen, Frederiksberg C, Denmark

The Brassicaceae family contains many agriculturally important crops such as cabbage, broccoli, and oilseed. This family has evolved numerous adaptations that defend against herbivores and pathogens, including a unique weapon, the ‘glucosinolate-myrosinase’ system. During predation, unnatural cell breakage releases myrosinase enzymes that hydrolyze glucosinolates from damaged plant tissues, yielding products that are toxic and/or that can be growth-inhibiting to animals. Glucosinolates, including hundreds of diverse thioglucosides, are mobile secondary metabolites synthesized, transported and stored among different plant tissues and organs. In order to prevent the hydrolysis of glucosinolates by myrosinases in intact tissues without predation, myrosinases are supposed to be separated from glucosinolates via distinct cellular or subcellular compartments. In growing plant shoots, myrosinases are synthesized and stored only in two types of myrosin cells (MCs), stomata and large cells located near the phloem termed myrosin idioblasts. These two cell types originate from different stem cells, although they harbour a shared developmental mechanism. Although the proximity of glucosinolates and MCs has been widely recognized to facilitate cell disruption thereby discouraging predation while minimizing metabolic costs, the spatial relationship between myrosinases and glucosinolates is still under debate. Here we investigate the cellular localization of glucosinolate synthesis, transport and accumulation with respect to developing and mature MCs via co-localization of the promoter-reporter fusion of glucosinolate biosynthesis and MC-specific genes. The major glucosinolate compounds from mutants with decreased or increased numbers of MCs were measured by liquid chromatography-mass spectrometry. The data obtained strongly support a developmental separation between myrosin cells and the presence of glucosinolates.

Meng Li [email protected]

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DV.3: Stone cells and the conifer defense syndrome against insects

Justin G. A. Whitehill1, Macaire Man Saint Yuen 1, Hannah Henderson1, Mathias Schuetz2, Jennifer Bryan1,3, Barry Jacquish4, Ward Strong4, A. Lacey Samuels2, Shawn D. Mansfield5, and Joerg Bohlmann1,2,6

1Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC, Canada V6T 1Z4 2Department of Botany, University of British Columbia, 6270 University Boulevard, Vancouver, BC, Canada V6T 1Z4 3Department of Statistics, University of British Columbia, Vancouver, BC, Canada 4British Columbia Ministry of Forests, Lands, and Natural Resource Operations, Victoria, BC, Canada V8W 9C2 5Department of Wood Science, University of British Columbia, 2424 Main Mall, Vancouver, BC, Canada V6T 1Z46Department of Forest and Conservation Sciences, University of British Columbia, 2424 Main Mall, Vancouver, BC, Canada V6T 1Z4

Conifers display an array of physical and chemical defenses against insects. Stone cells have extremely thickened cell walls that provide a robust physical defense associated with resistance against bark beetles and weevils. Abundance of stone cells in Sitka spruce (Picea sitchensis) cortex is positively correlated with resistance to the white pine weevil (Pissodes strobi). However, the mode of action by which stone cells interfere with growth and development of weevil larvae is unknown. We provide a detailed characterization of the stone cell phenotype. Contrasting Sitka spruce genotypes resistant (R) or susceptible (S) to the weevil revealed differences in stone cell distribution, abundance, morphology, biochemical composition, and molecular characteristics. Weevil larvae that fed on R trees with high quantities of cortical stone cells were significantly delayed developmentally. Changes to terpene profiles were observed for R trees but absent in S trees. Transcriptomes of R and S trees also revealed major and significant differences prior to and upon feeding by larvae. Stone cells hindered establishment of neonate larvae and impacted mandibles and relative growth rates of third instar larvae in bioassays. We conclude that stone cells interfere with the completion of the insect lifecycle and enhance the defensive capabilities of additional endogenous chemical and physical traits. Putative NAC transcription factors controlling stone cell biogenesis in R spruce have been identified and are being evaluated using transgenic Arabidopsis, tobacco and spruce. Identification of genes involved in stone cell formation will facilitate the production of weevil resistant spruce through transgenic and conventional breeding approaches.

Justin G. A. Whitehill – [email protected]

DV.4: Controlled alteration of pattern and extent of veins in monocot species Jim MATTSSON, Shawna WICKERMANN, Fatemeh ABEDI-SAMAKOUSH, Nicole SZULC Department of Biological Sciences, Simon Fraser University, Burnaby, BC There is an interest in increasing photosynthetic yield in rice and wheat growing in hot and dry climes by equipping them with traits typical of plants using C4 photosynthesis. Since C4 photosynthesis occurs primarily in bundle sheath cells of veins, C4 species have more veins than C3 species. While it is well known that the hormone auxin and its polar transport play key roles in the patterning of the pinnate venation in eudicot plants such as Arabidopsis, currently published evidence suggest minor roles in the patterning of the striate venation in monocot plants. In this study, we assessed the role of auxin in monocot leaf vein patterning. We found that chemically induced auxin transport inhibition of growing monocot plants dramatically increased the number of veins in chives and onion, but resulted in a limited increase of vein number in wheat and ryegrass and had no effect on vein number in rice plants. Since it is known that inhibition of auxin signaling in Arabidopsis results in leaves with reduced number of veins, we tested an auxin signaling inhibitor on rice, and found that it results in a reduced number of veins. Similarly, a mutant defective in an Auxin Response Factor-encoding gene also showed a reduced number of leaf veins. In summary, these results suggest that, although with clear differences, auxin transport and signaling acts in the regulation of vein formation also in monocot species, providing an opening for genetic manipulation of vein number towards the development of C4 rice and wheat. Jim Mattsson [email protected]

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DV.5: Redox regulation of after-ripening induced wheat seed dormancy release revealed through tissue-specific proteomics and antioxidant capacity profiling

Junjie HU-SKRZENTA1,2*, Nataša RADOVANOVIC1, Michelle L. RAMPITSCH1, Abir U. IGAMBERDIEV2, Natalia V. BYKOVA1

1. Morden Research and Development Centre, Agriculture and Agri-Food Canada, Morden, MB 2. Department of Biology, Memorial University of Newfoundland, St. John's, NL Alleviation of wheat seed dormancy during dry after-ripening involves reactive oxygen species (ROS) production, which triggers redox regulation of protein thiols and changes in functional proteome. Seed dormancy is closely associated with pre-harvest sprouting (PHS) tolerance. Tissue-specific redox proteomics studies contribute to understanding of molecular mechanisms controlling dormancy and after-ripening and further reveal candidate genes likely to contribute to PHS resistance traits in wheat. We analyzed dormancy genotype-associated and after-ripening induced alterations in embryo and aleurone redox proteomes using hard white spring wheat hybrid doubled haploid lines with marginal dormancy phenotypes, fluorescent labeling of redox active cysteines, high resolution 2-‐DE, LC-MS/MS analysis in conjunction with wheat EST database mining, quantitative spectral counting analysis, bioinformatics interpretation and visualization through MapMan. Redox proteomic analysis resulted in 689 high confidence protein identifications with at least two-fold change in protein expression and/or redox level between dormant and after-ripened seeds, from which 115 and 204 proteins showed significant disulfide/thiol ratio changes in dormant embryo and aleurone, respectively. Diagnostic proteins in dormant embryo were found to be involved in ascorbate regeneration, membrane trafficking, DNA methylation-dependent gene silencing, ABA and stress-responsive regulation of reproductive development. The level of total glutathione was significantly higher in dormant than in non-dormant and after-ripened embryos, whereas the level of total ascorbate increased 2-3 fold upon after-ripening. Therefore, glutathione and ascorbate antioxidants have different roles in redox control of seed dormancy and germination potential.

Junjie Hu-Skrzenta, [email protected]

DV.6: Translating frost tolerant seed degreening from Arabidopsis to Canola Mendel Perkins1,2, Logan Skori1*, Muhammad Jamshed1, Subramanian Sankaranarayanan1, Marcus Samuel1 1. Department of Biological Sciences, The University of Calgary, Calgary, AB 2. University of British Columbia, Department of Botany, Vancouver, BC Non-lethal frost during critical stages of Brassica napus (Canola) embryo development is known to significantly increase seed chlorophyll levels. Increasing seed chlorophyll levels result in seed downgrading, which results in economic penalties to producers. Components of the seed de-greening pathway have been identified in the model organism Arabidopsis thaliana. ABI3, a transcription factor involved in mediating abscisic acid (ABA) responses has been shown to regulate a large suite of genes related to seed maturation and de-greening. During seed maturation, ABI3 is required for the transcriptional activation of SGR2, which encodes a magnesium dechelatase implicated in both chlorophyll and photosystem degradation. ABI3 has been shown to interact with the SGR2 promoter to drive transcriptional activity. In Arabidopsis, overexpression of ABI3 was sufficient to impart frost-tolerant seed degreening. Given the high level of sequence similarity of SGR2 and ABI3 between species, it is expected this technology can be translated into Canola. To accomplish this, Brassica napus homologs of SGR2 and ABI3 were isolated and tested for their ability to perform similar functions using appropriate Arabidopsis mutants. In parallel, transgenic Canola lines constitutively overexpressing ABI3 were developed. These transgenic lines can de-green more completely than wild-type lines following frost exposure. Despite being expressed ectopically, the downstream up-regulation of SGR2 was restricted to the seed suggesting additional regulation in the leaves prevents ABI3 overexpression from initiating premature leaf chlorophyll catabolism. In addition, estimations of the transgenic lines growth characteristics and yield suggest an absence of deleterious pleiotropic effects. Logan Skori [email protected]

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Biochemistry Thursday July 6 3:20-4:50pm; FSC Room 1001 Chair: Greg Moorhead, University of Calgary BC.1: Activation of mitochondrial protein phosphatase SLP2 by MIA40 regulates seed germination R. Glen Uhrig1,2, Anne-Marie Labandera1 and Greg B.G. Moorhead1 1Department of Biological Sciences, University of Calgary, T2N 1N4 Calgary, Alberta, Canada 2Plant Biotechnology, Department of Biology, Swiss Federal Institute of Technology (ETH), Zurich, Switzerland Reversible protein phosphorylation catalyzed by protein kinases and phosphatases represents the most prolific and well-characterized post-translational modification known. Here, we demonstrate that Arabidopsis thaliana Shewanella-like protein phosphatase 2 (AtSLP2) is a bona-fide serine/threonine protein phosphatase that is targeted to the mitochondrial intermembrane space (IMS) where it interacts with the mitochondrial oxidoreductase ‘import and assembly protein 40’ (AtMIA40), forming a protein complex. Interaction with AtMIA40 is necessary for the phosphatase activity of AtSLP2 and is dependent on the formation of disulphide bridges on AtSLP2. Furthermore, by utilizing atslp2 null mutant, AtSLP2 complemented and AtSLP2 over-expressing plants, we identify a function for the AtSLP2-AtMIA40 complex in negatively regulating gibberellic acid related processes during seed germination. Results presented here characterize the first mitochondrial intermembrane space-localized protein phosphatase identified in photosynthetic eukaryotes as well as the first protein phosphatase target of the highly conserved eukaryotic MIA40 IMS oxidoreductase. Greg B.G. Moorhead [email protected] BC.2: Expression and properties of the mitochondrial and cytosolic forms of fumarase in germinating seeds of sunflower and maize Alexander T. EPRINTSEV1, Dmitry N. FEDORIN1, Oksana V. SAZONOVA1, Johanna Ludmila CETRE THYME1, Abir U. IGAMBERDIEV2* 1. Department of Biochemistry and Cell Physiology, Voronezh State University, 394018 Voronezh, Russia 2. Department of Biology, Memorial University of Newfoundland, St. John’s, NL A1B 3X9, Canada Fumarase (EC 4.2.1.2) catalyzes reversible interconversion of malate and fumarate. It operates in the tricarboxylic acid (TCA) cycle in mitochondria, and recently the cytosolic form has been detected. We investigated the expression of two fumarase genes (Fum1 and Fum2) and the activities of the mitochondrial and cytosolic isoforms of fumarase in sunflower (Helianthus annuus L.) cotyledons and maize (Zea mays L.) scutellum during germination. The mitochondrial and cytosolic forms of fumarase were purified to electrophoretic homogeneity from both plants. Both forms were tetrameric having similar molecular weight (~200 kDa) but differed in pH optimum (lower for the cytosolic form) and in affinity to malate. The cytosolic form was activated by Mg2+ and Mn2+, while the mitochondrial form was only moderately activated by Mg2+ and Mn2+ was not efficient. The highest fumarase activity and a high expression of the gene encoding the cytosolic form (Fum2) were observed during the period of maximum activity of the glyoxylate cycle enzyme isocitrate lyase. We suggest that the function of cytosolic fumarase in sunflower cotyledons and maize scutellum can be related to metabolism of succinate formed in the glyoxylate cycle. In the leaves of both investigated plants, the activity of fumarase was detected only in mitochondria and only one fumarase gene (Fum1) was markedly expressed, which indicates its participation mostly in the TCA cycle. This study, therefore, provides evidence of the dual localized function of fumarase in sunflower and maize in the course of mobilization of stored fats during seed germination. Abir Igamberdiev [email protected]

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BC.3: Peeling off the secrets of poplar cuticles Eliana GONZALES-VIGIL1, Charles A. Hefer2, Michelle E. von Loessl1, Jonathan La Mantia3, and Shawn D. Mansfield1. 1. Department of Wood Science, University of British Columbia, Vancouver, British Columbia, V6T 1Z4 Canada. 2. Biotechnology Platform, Agricultural Research Council, Private Bag X05, Onderstepoort, Pretoria, 0110, South Africa. 3. United States Department of Agriculture-Agricultural Research Service (USDA-ARS), Corn and Soybean Research, Wooster, OH 44691, USA. The cuticle is the extracellular hydrophobic barrier that protects the aerial surfaces of land plants; its outermost layer consists of cuticular waxes whose composition varies among species, tissues and developmental stages. A screen of natural accessions of Populus trichocarpa revealed that the leaf cuticular waxes are predominantly composed of alkenes. Alkenes are linear hydrocarbons with one or more double bonds, and despite their many industrial applications, including advanced biofuels, their biosynthetic origin in plants was completely unknown. Interestingly, in Populus, the accumulation of alkenes appears to be controlled by leaf development and is limited to the abaxial side of the leaf. By mining the leaf transcriptome of P. trichocarpa, a ®-ketoacyl CoA synthase (PotriKCS1) was subsequently found to be downregulated in leaves originating from accessions lacking alkenes. We demonstrate biochemically that PotriKCS1 elongates unsaturated fatty acids and is part of an eight-gene cluster of recent evolution in the Salicaceae; one that diversified the complement of cuticular lipids found in Populus trichocarpa leaves. Moreover, the accumulation of alkenes in the leaf cuticle was found to be associated with increased growth and resistance to the leaf spot pathogen Septoria musiva. This knowledge can be used to breed trees more resistant to diseases and better growth, or engineer plant oils in other species. Eliana Gonzales-Vigil [email protected] BC.4: Allosteric regulation of Brassica napus diacylglycerol acyltransferase 1 Kristian Mark P. CALDO1,2*, Jeella Z. ACEDO3, Rashmi PANIGRAHI2, John C. VEDERAS3, M. Joanne LEMIEUX2, Randall J. WESELAKE1 1. Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, Alberta, Canada 2. Department of Biochemistry, University of Alberta, Edmonton, Alberta, Canada 3. Department of Chemistry, University of Alberta, Edmonton, Alberta, Canada Diacylglycerol acyltransferase 1 (DGAT1) is a multi-pass membrane protein catalyzing the acyl-CoA-dependent biosynthesis of triacylglycerol. Previous metabolic control analysis showed that DGAT1 appears to modulate the flux of carbon towards oil accumulation in some organisms including canola (Brassica napus). Hence, DGAT1 has been extensively used as a molecular tool to boost oil content in plants, algae and microorganisms. Although plant and mammalian DGAT1 have been studied for several decades, the mechanisms of enzyme catalysis and regulation remain unclear. In this study, we showed that the hydrophilic N-terminal region of B. napus DGAT1 (BnaDGAT1) corresponds to a regulatory domain that is not necessary for catalysis. Biophysical analysis showed that this domain is composed of a disordered segment and a folded portion. The acyl donor substrate, acyl-CoA, regulates BnaDGAT1 activity through positive cooperativity by binding to an allosteric site within the folded portion. The disordered segment may have an autoinhibitory motif and a dimerization interface. Coenzyme A (CoA) was also identified as a non-competitive feedback inhibitor, the level of which may accumulate under limiting acyl-CoA conditions. The acyl-CoA binding site in the folded region also interacted with CoA. Furthermore, phosphatidic acid (PA), an intermediate in the Kennedy pathway, appeared to serve as a feedforward activator of BnaDGAT1. PA may have facilitated the conformational shift into the more active state and provide relief to autoinhibition. Based on these structural and kinetic analyses, an enzyme model is presented demonstrating the effect of the identified allosteric effectors on DGAT1 catalysis. Kristian Mark P. Caldo [email protected]

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BC.5: A pathway for disposal of 5′-deoxyadenosine, a toxic byproduct of radical SAM enzymes Guillaume A.W. Beaudoin1, Andrew D. Hanson1 1. Horticultural Sciences Department, University of Florida, Gainesville, FL, USA Radical S-adenosylmethionine (SAM) enzymes, essential in all domains of life, produce the toxic by-product 5′-deoxyadenosine. In mammals, 5′-deoxyadenosine is converted to 5-deoxyribose, which is reduced and excreted. The fate of 5-deoxyribose in other organisms is unknown. We recently uncovered a pathway for 5-deoxyribose disposal in bacteria. Diverse taxa have a three gene cluster encoding paralogs of the kinase and isomerase from methionine salvage, and a fuculose 1-phosphate aldolase-like enzyme. This trio of recombinant enzymes from Bacillus thuringiensis converted 5-deoxyribose to dihydroxyacetone phosphate and acetaldehyde via 5-deoxyribose 1-phosphate and 5-deoxyribulose 1-phosphate. This type of pathway could potentially operate in plants, taking advantage of the promiscuity of the methionine salvage enzymes 5-methylthioribose kinase and 5-methylthioribose 1-phosphate isomerase (i.e. their respective abilities to act on 5-deoxyribose and its 1-phosphate ester). In this scenario the only missing enzyme would be an aldolase, for which certain candidates can already be proposed. Guillaume Beaudoin [email protected]

BC.6: Cell-type-specific metabolism: Biosynthesis and composition of cuticular waxes covering Arabidopsis trichomes

Daniela HEGEBARTH1, Lukas BUSTA2, Reinhard JETTER1,2 1. Department of Botany, University of British Columbia, BC 2. Department of Chemistry, University of British Columbia, BC

To protect plants against biotic and abiotic stress, the waxy cuticle must coat all epidermis cells. We have used several independent approaches to test whether cell-type-specific differences exist between wax compositions on trichomes and other epidermal cells of Arabidopsis thaliana: (i) the leaf wax composition was monitored during leaf expansion, which results in a steady decrease in the trichome:pavement ratio; (ii) the total waxes from a mutant lacking trichomes (gl1) were compared to waxes from wild type and a trichome-rich mutant (cpc tcl1 etc1 etc3); (iii) leaf trichomes were isolated and their waxes were analyzed. All three experiments showed that trichome wax comprised fewer classes of wax compounds than adjacent pavement cells. Most interestingly, trichome wax contained relatively high concentrations of C32+ compounds, suggesting differences in the elongation machineries of different epidermis cell types. This finding led us to identify the ketoacyl-CoA synthase KCS16 as a crucial condensing enzyme for fatty acyl chain elongation beyond C32. Taken together, our results show that Arabidopsis leaf trichomes have autonomous wax biosynthesis machinery generating a distinct surface wax mixture, likely adapted to specific functional needs of these exposed epidermis cells with extreme surface-to-volume ratios.

Reinhard Jetter: [email protected]

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Applied Plant Biology Thursday July 6 3:20-5:05pm; FSC Room 1005 (7 Talks) Chairs: Diane Edwards, ABI Environmental Services Ltd & Samir Debnath, AAFC, St John’s AP.1: Conservation of Plant Biodiversity: The role of in vitro technologies Praveen K. Saxena1 1. Gosling Research Institute for Plant Preservation (GRIPP), Department of Plant Agriculture, University of Guelph, Guelph, ON, Canada N1G 2W1

Current climate change patterns, in combination with human activity and the spread of diseases, are leading to a global mass extinction of both ecologically and economically important species. Unfortunately, existing approaches for addressing these anticipated challenges are insufficient, creating an urgent need for researchers to develop new and effective conservation models and technologies. In vitro technologies provide an efficient, cost-effective approach for rapid plant multiplication and long-term conservation of plant germplasm. At the Gosling Research Institute for Plant Preservation (GRIPP), our mandate is to preserve critically endangered species as well as agriculturally important plant genetic resources through development and application of in vitro technologies for micropropagation and cryopreservation in order to enhance the resilience and biosecurity of plants. This presentation will provide an overview of GRIPPs most recent efforts for conserving several Canadian species including American elm, Cherry birch, Saint John’s wort, Hill’s thistle, and Golden paint brush in order to demonstrate the importance and application of in vitro technologies for long-term conservation of endangered and cultivated plant genetic resources. These examples will also highlight the urgent need for research in areas related to in vitro morphogenesis, as greater research in this area can lead to the development of new and innovative conservation strategies. [email protected]

AP.2: Morphogenesis and Regeneration of Breadfruit (Artocarpus altilis): A Staple Crop for Food Security

Susan MURCH1*

1. Chemistry, University of British Columbia, Kelowna, British Columbia, Canada, V1V 1V7

Breadfruit (Artocarpus altilis (Parkinson) Fosberg) is a traditional staple crop of the Pacific listed as a priority food crop on Annex 1 of the International Treaty on Plant Genetic Resources for Food and Agriculture. Distribution of breadfruit trees was limited by difficulties in propagation. The objective of this program was to develop in vitro mass propagation and ex vitro acclimatization methods to provide breadfruit trees for food security projects in the tropics. Methods were optimized for shipment of breadfruit shoots from tropical field sites and control of fungi and bacteria contamination on field harvested breadfruit tissues. The production of phenolics that damage explants was controlled with a phenylalanine ammonia lyase inhibitor. Breadfruit explants contain regenerative meristematic cells in the interior pith of woody shoots that was stimulated to divide and produce callus by an induction medium containing zinc pyrithione, kinetin, benzylaminopurine, gibberellic acid, glutamine, and 2-aminoindane-2-phosphonic acid. Different cultivars required different combinations of phytohormones for induction of regeneration including cytokinins, auxins and salicylic acid. In vitro grown plantlets were developed from de novo shoots on a media containing indolebutyric acid or a basal media devoid of plant growth regulators. The tissue cultured plants were screened for known pathogens, acclimatized to greenhouse conditions and distributed to food security projects. To date, more than 100,000 trees have been planted in 45 tropical countries. The mass propagation of breadfruit provides a crop that can be grown in sustainable agro-food-forests resilient to climate change to produce nutritious food products for world markets.

Susan Murch [email protected]

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AP.3: 36 years of breeding---novel low-light tolerant (LLT PlantsTM) ‘First Canadian’ lemon, ‘First Canadian Golden’ lime, and other home windowsill vegetables for northern regions

M.P.M. NAIR1 and Karen TANINO2

1. 4461 Clarence Ave. South, Grasswood, SK Canada S7T 1A7;

2. Department of Plant Sciences, College of Agriculture and Bioresources, Univ. of Saskatchewan, Saskatoon, Canada S7N 5A8.

Thirty-six years ago, shade-adapted tropical citrus (bush type) from South India was brought to Saskatchewan and used as the female parent for breeding with U.S. commercial cultivars. After three generations of crossing citrus with multiple blooming cycles, the new lines showed excellent performance as a dwarf plant in the home windowsill environment in Saskatchewan. These citrus plants will flower and fruit year-round even under east windowsill conditions (average irradiance 120 µmolm2s1 ). Some citrus plants are producing up to 12 – 15 commercial size lemon fruit/plant in a 15-cm pot. Quality traits include seedlessness, thin-exocarp, juicy, Brix = 7. By contrast, currently available Citrus plants in the marketplace are limited to outdoor high light, frost-free conditions of the southern citrus belt. The concept of home “Windowsill Farming” is the basis of a rapidly growing Garden Centre and Greenhouse industry to supply low light adapted crops. With increasing urbanization and high rises, decreasing access to land, increasing air pollution, cloud/rain in some regions, along with high interest in food security, the demand for such plants is significant. Furthermore, during the winter in temperate climates where homes are heated with fossil fuels, the CO2 levels can be extremely high. Plants can reduce our own carbon footprint, improve air quality while producing food. Low light tolerant crops may also be grown under canopies thereby producing two greenhouse crops in the same space. The purpose of this presentation is to introduce this novel germplasm, concept and techniques of growing low light tolerant fruits and vegetables.

M.P.M. Nair [email protected]

AP.4: Potato greening: Towards developing potato clones tolerant to greening Bourlaye FOFANA1, Ashok SOMALRAJU1,2, Kaushik GHOSE1,3, David MAIN1

1. Charlottetown Research and Development Centre, Agriculture and Agri-Food Canada, 440 University Avenue, Charlottetown, Prince Edward Island, C1A 4N6, Canada

2. University of Prince Edward Island, 550 University Avenue, Charlottetown, Prince Edward Island, C1A 4P3, Canada

3. Current address: Department of Plant and Soil Science, Texas Tech University, Food Technology Building, 2802 15th Street, Lubbock, TX 79409-2122, United States

Potato and its processed products are major components in modern diets. However, 2-3% of potato crops are lost at the farm gate and during postharvest storage due to undesirable tuber greening following light exposure. Tuber greening involves the chlorophyll synthesis in the cortical parenchyma cells beneath the periderm and a simultaneous accumulation of toxic glycoalkaloid compounds in the light-exposed area. Toxic glycoalkaloids are a major concern to human health and greened potatoes are discriminated against by the consumers. Thus, elimination of this undesirable trait may involve the alteration of more than one biosynthetic pathway. We have developed and characterized an EMS mutagenized diploid potato population with the aim of identifying potato lines low in anti-nutritional factors. This talk will focus on the development paths of the tolerant lines as well as some of the molecular and physiological processes behind greening tolerance.

Bourlaye Fofana [email protected]

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AP.5: Effect of varieties and cooking methods on starch digestibility and glycemic impact of early potatoes

Reena Grittle PINHERO1, James A. SULLIVAN2, Qiang LIU3, Rong CAO3, Massimo MARCONE1, Rickey Y. YADA4 1 Department of Food Science, University of Guelph, Guelph, Ontario 2 Department of Plant Agriculture, University of Guelph, Guelph, Ontario 3 Agriculture and Agri-Food Canada, Guelph Food Research Centre, Guelph, Ontario 4 Faculty of Land and Food Systems, University of British Columbia, Vancouver Potatoes, the number one vegetable crop in the world, a staple in many diets, offers one of the best nutritional value on a cost/weight basis. The objective of this study was to identify potato varieties, from eight varieties, with nutritional benefits on the basis of starch profiles and the effect of cooking methods. Cooking methods included retrogradation (boiling followed by refrigeration at 4°C), retrogradation and reheating in the microwave, baking and microwaving. Variety and cooking methods significantly affected the starch profile. Significantly lowest total starch and amylose contents were obtained in the variety Smart. Purple Fiesta had the highest total starch and amylose contents. Smart registered the lowest rapidly digestible starch irrespective of cooking methods. Only baked and microwaved potatoes showed significant differences in resistant starch among the cooking methods. Among baked potatoes, highest resistant starch was obtained in Purple Fiesta and French Fingerlings whereas Red Thumb and Smart registered the highest values in microwaved potatoes. Estimated glycemic index (eGI) of varieties ranged from low to medium (48-66) in various cooking methods. Lowest eGI was obtained in Smart. In general, retrograded method registered the lowest eGI and glycemic load (GL) in all varieties. GL of all the varieties and cooking methods tested fell under low GL and ranged from 3 to 10. Smart registered the lowest eGI and GL among all the processing methods. Based on starch profile and glycemic impact, Smart was identified as having the best nutritional quality. Reena Grittle Pinhero [email protected] AP.6: Biotic constraints for leafy vegetable production in Southeast Asia: Disease survey, pathogen characterization, and screening for disease resistance

R. R. BURLAKOTI1,5*, J.-R. CHEN1, F.-I. HO1, Z.-M. SHEU1, J.-F. WANG1, C.H. NGUYEN2, S. KHODSIMOUANG3, and H.C. HY4 1 World Vegetable Center (WorldVeg), Shanhua, Tainan 74199 Taiwan 2 Vietnam Academy of Agricultural Science, Hanoi, Vietnam 3 Department of Agriculture, Vientiane, Lao PDR 4 General Directorate of Agriculture, Phnom Penh, Cambodia 5 Current addresses: Agassiz Research and Development Center, Agriculture and Agri-Food Canada, Agassiz, BC

Leafy mustard (Brassica juncea), pak-choi (B. rapa var. chinensis), choi sum (B. rapa var. parachinensis) etc. are important vegetables crops in Vietnam, Cambodia and Lao PDR. Survey and characterizations of the major diseases have not been conducted in these countries. The objectives of the research were: (i) to identify major diseases and detect seed-borne pathogens in from leafy vegetables, (ii) to develop suitable screening methods and identify resistant sources for major bacterial and fungal diseases. Disease surveys during 2014 to 2016 revealed that root rot and web blight caused by Rhizoctonia solani was the most destructive fungal disease (up to 50-60 % disease incidences). Other fungal diseases present at low to moderate levels were: downy mildew (Peronospora parasitica), damping off (Pythium spp., Fusarium spp.), Alternaria leaf spot (Alternaria spp.) and other leaf spots. Black rot caused by Xanthomonas campestris pv. campestris (Xcc) was the most destructive bacterial disease in all three countries. Evaluation of a total of 85 seed samples collected from three countries in 2014 and 2016 detected Xcc contamination only in 3 seedlots from Lao PDR in 2014, whereas one seedlot from Vietnam were contaminated with Xcc in 2016. Several seedlots from all three countries were contaminated with Alternaria spp. (up to 70% contamination in Alternaria specific media). Evaluation of 50 accessions of leafy mustards from WorldVeg’s genebank showed none of accessions had good level of resistance to Rhizoctonia damping off and black rot caused by Xcc, which indicates the need of more efforts to identify the sources for resistance.

* Rishi Burlakoti [email protected]

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AP.7 Preharvest treatment application: A new era for UV-C hormesis Marie Thérèse CHARLES1, Yanqun XU 1,2, Zhichun XIE1,3, Joseph ARUL4, Zisheng LUO2. 1. Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, Saint-Jean-sur-Richelieu, QC, J3B 3E6, Canada 2. College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310058, People’s Republic of China 3. College of Forestry, Northwest A&F University, Yangling, Shaanxi 712100, China 4. Horticultural Research Center and Department of Food Science and Nutrition, Université Laval, Sainte-Foy, Québec, Canada G1K 7P4 Hormesis is the phenomenon that underlies a set of favorable responses observed in a biological system subjected to low levels of exposure to stressors. Over the last thirty years, hormetic levels of short-wave ultraviolet (UV-C) radiations, applied at the post-harvest stage, have been studied for their potential to extend shelf life and improve quality of a wide range of horticultural products. UV-C hormesis has been found to be effective in delaying senescence, inactivating pathogenic and deterioration causing microorganisms, preventing losses due to rot and disease development, and improving the bioactive components of the products. Unfortunately, in the case of fragile commodities, the non-systemic action of UV-C slows down the adoption of this practice by the fresh fruit and vegetable (F & V) industry. Recent considerations have been given to F & V responses to UV-C hormesis when applied at the prehavest stage. Early findings highlighted that this new approach could be considered in pest management and as a sustainable way to improve fresh F & V quality and composition. [email protected]

Biotic Interactions Thursday July 6 3:20-4:50pm; FSC Room 1003 Chair: Cara Haney, University of British Columbia BI.1: Manipulating the crosstalk plant response to biotic and abiotic stimuli in horticultural plants Valérie GRAVEL1, Varinder SIDHU1 1. Department of Plant Science, McGill University, Sainte-Anne-de-Bellevue, QC The great demand for high-quality plant produce with elevated concentrations of health-beneficial compounds has created a need for the agricultural industry to investigate underlying processes leading to the accumulation of such compounds. In sustainable agricultural production systems, persistent abiotic stresses (linked to temperature, photoperiod or water and nutrient deficiencies) or biotic stresses (triggered by soil microbial communities) are known to promote the accumulation of health-beneficial compounds such as flavonols and anthocyanins. Although accumulation of specific compounds in response to specific stresses is well documented, a thorough understanding of the patterns of plant response to the signalling interaction resulting from simultaneous abiotic and biotic stress is still sorely lacking. The effect of biotic and abiotic stimuli on plant development and fruit quality was therefore evaluated using strawberry (Fragaria vesca and Fragaria x ananassa) as a model plant. More specifically, manipulating plant response through adapted photoperiods, specific light quality and temperatures resulted in an improved flower bud induction and overall plant architecture. Combined with a higher soil microbial activity typical of sustainable production systems, higher fruit yields and health-beneficial compound accumulation was also observed leading the way to develop integrated and effective management practices. [email protected]

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BI.2: RNA interference two ways: molecular fungicides and durable plants to control Sclerotinia sclerotiorum A. G. MCLOUGHLIN, S. WHYARD, M. F. BELMONTE Department of Biological Sciences, 50 Sifton Road, University of Manitoba, MB, R3T 2N2, Canada Infecting over 450 plant species, sclerotinia stem rot (white mold, Sclerotinia sclerotiorum) substantially threatens Canadian canola, which contributes over 26 billion dollars to the economy. Traditionally, Sclerotinia control involved the use of broad-spectrum fungicides, which are neither economical or effective. Crop rotations, which are also used, fail due to the promiscuous host range of Sclerotinia and the formation of durable, melanized resting structures called sclerotia, which persist in the soil for up to 10 years. Consequently, there is an urgent need for novel species-specific methods to mitigate Sclerotinia. Our novel strategy exploits the inherent cellular defense process known as RNA interference (RNAi) by employing in vitro or in planta synthesized double stranded RNAs (dsRNA). Upon encountering the designer molecules, the cell processes the dsRNA to specifically target homologous transcripts. Using a comprehensive bioinformatics pipeline, Sclerotinia genes were identified and Sclerotinia-specific dsRNA molecules were synthesized. Target gene knockdown was confirmed and quantified using quantitative real-time PCR from RNA isolated from fungal liquid cultures. Using a petal inoculation method that mimicked aggressive infection conditions, over 70 dsRNA molecules were evaluated. Lesion size was significantly reduced on mature leaf tissues by up to 85%. To protect plants throughout their lifecycle, we developed constitutively expressing dsRNA expressing Arabidopsis thaliana. When challenged with Sclerotinia, transgenic RNAi plants reduced lesion size up 75% when compared to wild-type controls. Taken together, we have developed two novel solutions to combat this devastating fungus: a species-specific molecular fungicide capable of controlling fungal infection on the leaf surface and transgenic RNAi plants protected throughout the lifecycle. Mark Belmonte [email protected] BI.3: Can pine germinants supply nitrogen to their ectomycorrhizal fungal partners? Josh M. SMITH1, Matthew D. WHITESIDE1,2 and Melanie D. JONES1 1. Biology Department, University of British Columbia Okanagan Campus, Kelowna, British Columbia V1V 1V7

Canada 2. Current Address: Department of Ecological Sciences, Vrije Universiteit, Amsterdam, The Netherlands Ectomycorrhizae are ancient symbiotic relationships between fungi and plant roots. One of the main benefits to plants is that ectomycorrhizal fungi translocate soil nitrogen to roots. In this study, we found evidence for the reverse: movement of nitrogen from the plant toward the fungus. We grew Pinus contorta seedlings in association with Suillus tomentosus in microcosms where most of the nitrogen was located in a small well accessible only by hyphae. Twelve weeks after initial nutrient placement in the wells, they were replenished with an organic (glycine) or inorganic (NH4) nitrogen source. Before well replenishment N concentration of shoots and roots ranged from 36.6 ± 0.9 to 44.9 ± 8.2 (µg N mg-1, ± SD) but 48 hrs after the hyphal nitrogen was applied, tissue N concentrations dropped to 11.2 ± 1.5 to 13.5 ± 2.1 µg N mg-1. If hyphae entering the well were severed prior to N addition, or if water was added to the wells, no loss of N from seedlings was observed. We speculate that, after a period of nutrient starvation, exposure of the fungus to a renewed nutrient source induced a priming effect by the host plant. The N translocated from the plant may have facilitated fungal exploration of the new nutrient source. Further research into bi-directional nutrient exchange between mycorrhizal symbionts is required to determine the frequency and conditions under which this host-to-fungus nutrient transfer occurs. Melanie Jones [email protected]

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BI.4: Molecular and Microscopic Dissection of Plant-Microbe Interactions in Early Land Plants Philip CARELLA1, Anna GOGLEVA1, Marta TOMASELLI1, Carolin ALFS1, and Sebastian SCHORNACK1 1. Sainsbury Laboratory, University of Cambridge, Cambridge, United Kingdom Fossil evidence dates the first occurrence of terrestrial plants to over 450 million years ago. Strikingly, several fossils demonstrate close physical associations between ancient land plants and filamentous microbes that are similar in nature to modern endosymbiotic structures such as arbuscules. Indeed, symbiotic interactions with arbuscular mycorrhizal fungi or other filamentous microbes have been documented in extant early land plants such as liverworts and hornworts, yet interactions with pathogens remain unstudied. To address this gap in knowledge, we characterized interactions between a broad host-range hemibiotrophic filamentous pathogen and several liverwort species. We establish that liverworts of the Lunularia and Marchantia genera are susceptible to colonization and succumb to disease. Microscopic analysis of these interactions demonstrates the presence of intracellular infection structures, which differ in morphology in different liverwort species. Further, we performed expression studies to characterize both host and pathogen transcriptional responses, identifying novel and conserved responses during colonization. Our results demonstrate that filamentous pathogens are well suited to manipulate and colonize early land plants. Moreover, we establish a key liverwort-microbe pathosystem that will be a valuable tool to address important questions pertaining to the evolution of defense and symbiosis programs during the colonization of land. Dr. Philip Carella [email protected] BI.5: A scalable comparative genomics platform for identifying plant-associated genes in Pseudomonas spp. Ryan A. Melnyk1, Sarzana S. Hossain1, and Cara H. Haney1 1. Department of Microbiology and Immunology, University of British Columbia, Vancouver, BC

High throughput sequencing has led to an expansion of genomes of well-studied organisms, with over 1000 genome sequences from the genus Pseudomonas alone. Pseudomonas spp. display many interesting plant-associated behaviors and have large and variable genomes, making this genus an excellent system for investigating relationships between bacterial genes and plant host interactions. However, conventional methods for detecting orthologs do not scale well to modern genome databases, as their computational complexity exponentially relative to the number of genomes. Therefore, we developed a pipeline that develops gene family models from a training dataset and “propagates” these models to additional genomes, scaling linearly. Applying our pipeline to the genus Pseudomonas, we identified 24,066 gene families that covered 98.0% of the input gene content and 92.0% of the “propagated” gene content. To demonstrate the PyParanoid platform, we investigated the genetic basis of a surprising plant-associated behavior. When the soil isolate P. fluorescens N2C3 is applied to the roots of Arabidopsis thaliana seedlings, development is inhibited, unlike close relatives that promote plant growth. Using the PyParanoid database, we compared the genome of N2C3 to close relatives and identified a genomic island containing genes for non-ribosomal peptide synthases and a quorum-sensing system. We disrupted the quorum-sensing system in N2C3 which restored seedling development but did not impair rhizosphere colonization. Further analysis showed that this island is similar to virulence genes in plant pathogenic Pseudomonas strains, suggesting horizontal transfer. This example shows the utility of genomics to inform understanding of plant-associated bacterial traits. Ryan A. Melnyk [email protected]

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BI.6: Horizontal transfer of microbes between seaweed neighbours Melissa Y. CHEN1, Laura PARFREY1 1. University of British Columbia, Vancouver, BC Seaweeds possess complex surface microbiota that are integral for their health [1][2]. For instance, they provide improved nutrient acquisition for vitamins like B12 [3] and promote immune defence against potential pathogens [4]. Thus, it is important to understand the many factors that control microbial community assembly on seaweeds. The water column is considered to be the main source of microbes for seaweed microbiota recruitment, and it has been shown that the presence of seaweeds can in turn affect the water column community [5], [6]. However, it is unknown whether seaweeds influence each other’s microbiota: that is, does the identity and proximity of nearby seaweeds influence the assembly of microbial communities on individual seaweed? To test this, we incubated meristematic fragments of Nereocystis luetkeana with combinations of other seaweeds: full blades of Nereocystis, Mastocarpus sp., or both. Then, we sampled both the water column community and swabbed the surface of all meristematic Nereocystis to see whether the microbial community composition changed in treatments where they were co-incubated with other seaweeds. What we find is that the microbial community associated with meristematic Nereocystis does change depending on its co-incubate. Therefore, seaweeds appear to not only influence the water column microbial community, but also other individuals in the nearby seaweed community. Ecologically, this experiment demonstrates the importance of considering community-wide dynamics in studying community assembly and hopefully highlights the interconnectivity between seaweed, its microbiota, and its neighbours. Melissa Chen [email protected]

References:

[1] S. Egan, T. Harder, C. Burke, P. Steinberg, S. Kjelleberg, and T. Thomas, “The seaweed holobiont: Understanding seaweed-bacteria interactions,” FEMS Microbiol. Rev., vol. 37, no. 3, pp. 462–476, 2013.

[2] T. Staufenberger, V. Thiel, J. Wiese, and J. F. Imhoff, “Phylogenetic analysis of bacteria associated with Laminaria saccharina,” FEMS Microbiol. Ecol., vol. 64, pp. 65–77, 2008.

[3] M. T. Croft, A. D. Lawrence, E. Raux-Deery, M. J. Warren, and A. G. Smith, “Algae acquire vitamin B12 through a symbiotic relationship with bacteria.,” Nature, vol. 438, pp. 90–93, 2005.

[4] F. C. Küpper, D. G. Müller, A. F. Peters, B. Kloareg, and P. Potin, “Oligoalginate recognition and oxidative burst play a key role in natural and induced resistance of sporophytes of Laminariales,” J. Chem. Ecol., vol. 28, no. 10, pp. 2057–2081, 2002.

[5] C. Lam and T. Harder, “Marine macroalgae affect abundance and community richness of bacterioplankton in close proximity,” J. Phycol., vol. 43, no. 5, pp. 874–881, 2007.

[6] C. Lam, A. Stang, and T. Harder, “Planktonic bacteria and fungi are selectively eliminated by exposure to marine macroalgae in close proximity,” FEMS Microbiol. Ecol., vol. 63, no. 3, pp. 283–291, 2008.

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Posters – Session 1 Wednesday July 5, FSC Atrium 11:20am -12:30pm

Abiotic Stress Posters (101-‐AB to 120-‐AB) 100-AB: A modeling approach: The effect of endogenous and exogenous chelating agents on metal uptake by Raphanus sativus L. cv. Crimson Giant. Sheila MACFIE, Sergio Ari DOMINGUEZ ROMERO* Department of Biology, University of Western Ontario, London, ON Metal contamination is a common environmental problem. One answer has been to apply exogenous chelators to contaminated soil so as to improve the efficiency of phytoremediation. However, the optimal amounts and types of chelators have not been determined. We took a geochemical modelling approach to estimate the effects of four synthetic chelators on the solubility of four metals and compared these predictions to the amounts of these metals taken up by radish (Raphanus raphanistrum subsp. sativus (L.) Domin). We used a hydroponic system to eliminate soil-metal chemistry and to get a clearer understanding of the direct metal-chelate interactions. As predicted by the model (Visual MINTEQ 3.1), iron solubility depended on the presence of a chelator and the solubility of cadmium, copper or zinc were high with or without an exogenous chelator. In general, the predictions were supported by the amounts of metals actually taken up by the plants. Two surprises that emerged were the high amount of cadmium and the relatively low amount of iron measured in the plant tissues. Despite the apparently low iron, the plants did not show symptoms of iron-deficiency but, not surprisingly, the plants given cadmium were clearly stressed. We also considered the effect on metal solubility and uptake of low molecular weight organic acids (LMWOAs) exuded from the roots. Despite having high concentrations of LMWOAs in solution, their effects on metal solubility were negligible. The model indicated that this was due to lack of metal-LMWOA complex formation. Sergio Ari Dominguez Romero [email protected]

101-AB: Role of RAE1 in Arabidopsis thaliana heat tolerance

Azizah ALHARTHY*, Charlene IBBETSON, Amy CLARK, Dana SCHROEDER Department of Biological Sciences, University of Manitoba, Winnipeg, MB Plants need to adapt to environmental conditions. Here we examine heat stress response in Arabidopsis thaliana. Studies in our lab have implicated DDB1 in heat tolerance. DDB1 interacts with DWD proteins, and six heat induced DWD genes were identified as candidate genes. Analysis of heat sensitivity in loss of function lines revealed that a RNA EXPORT1 (rae1) mutant allele resulted in heat sensitivity in adults and seedlings. We examined the effect of this allele on the RAE1 transcript and found that the transcript was still expressed in control conditions, however heat stress resulted in failure of intron 1 splicing. We examined stronger rae1 alleles, however were unable to identify homozygotes, suggesting gametic or zygotic lethality. Nonetheless heterozygotes of one of these stronger alleles also exhibit increased heat sensitivity. YFP-RAE1 was found to be nuclear localized, however following heat treatment it was also detected in cytoplasmic speckles. Our findings on the role of RAE1 in Arabidopsis heat tolerance will be presented. Azizah Alharthy [email protected]

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102-AB: Restoring Kelp Forest Habitat in the Salish Sea: Best Practices Braeden W.J. SCHILTROTH, Sherryl R. BISGROVE, William HEATH 1. Department of Biological Sciences, Simon Fraser University, Burnaby, BC Kelp beds are marine sanctuaries, providing some of the most productive ecosystems on the planet and serving as critical habitat and refuge for many species, including juvenile salmon. Recently, declines in kelp populations have been reported by several groups including fisherman and kelp harvesters in the Pacific Northwest. However, the extent and cause of this habitat loss is unknown. It is thought that an increase in stressors associated with climate change (rising ocean temperatures or acidification) is a major contributor. Thus, if lost habitats are to be restored, populations that are resilient to these stressors would be the best to use because they are most likely to survive. The objectives of this research are: 1) Identify kelp stocks capable of growing at sites with stressful conditions (high temperature) by correlating physical trends in the ocean with kelp distributions, 2) Evaluate whether early developmental stages of kelp populations from warm sites exhibit a higher tolerance to increased temperature than those from colder sites. 3) Assess restoration techniques and the performance of candidate kelp stocks in the lab. These studies will provide crucial data needed for habitat restoration efforts as they will allow for the selection of stress-resilient kelp stocks that are better adapted for survival in warmer oceans. Braeden Schiltroth [email protected]

103-AB: Effect of exogenous ABA and ABA mimic 1 (AM1) on the uptake and accumulation of zinc in grapevine exposed to excess zinc Changzheng SONG1,2, Simone D. CASTELLARIN1* 1. Wine Research Centre, The University of British Columbia, 2205 East Mall, Vancouver, BC V6T 1Z4, Canada 2. College of Enology, Northwest A&F University, Yangling 712100, Shaanxi, China Abscisic acid (ABA) is a stress phytohormone which can mitigate heavy metal toxicity. Zinc (Zn) contamination has been detected in soils of some wine regions. Exogenous ABA and ABA mimic 1 (AM1) were applied to study the influence on Zn uptake and accumulation in Vitis vinifera cv. Merlot seedlings exposed to excess Zn. The seedlings were treated with either normal or excess levels of Zn, and for two of the three treatments with excess Zn, 10 µM ABA or AM1 were applied. The results showed that root length of vines exposed to excess zinc notably decreased. The net photosynthetic rate was reduced by excess Zn at both 4 d and 10 d after treatment, while no significant alleviation effect was found on ABA or AM1 treated vines. As for photosynthetic pigments, evident decrease of Chl a was found between 4 d and 10 d after treatment with excess Zn, but ABA and AM1 alleviated the repressive effect. Histochemical analysis showed that more Zn was accumulated in root, trunk, stem, and petiole in excess Zn treatments at 10 d. Zn concentrations in root, trunk, and stem was significantly elevated in treatments with excess Zn at 10 d, and ABA alleviated the excess uptake in root. However, at 4 d after treatment, higher Zn concentration in root was found for ABA and AM1 treatments, indicating their transient promoting effect on root Zn uptake. Moreover, correlations of Zn concentration among tissues and interactions between Zn and other elements were found. [email protected]

104-AB: Investigating the role of the miR156-SPL network in heat stress response in Medicago sativa

Craig D. R. Matthews1,2*, Lisa Amyot2, David R. Smith1, Abdelali Hannoufa1,2

1. Department of Biology, University of Western Ontario, London, Ontario, Canada 2. London Research and Development Centre, Agriculture and Agri-Food Canada, London, Ontario, Canada

Extreme temperatures adversely affect plant development and, as a result, crop production. Heat stress affects cellular function and results in increased potential evapotranspiration. Heat stress induced changes to plant water status can be especially harmful when combined with drought. Alfalfa (Medicago sativa) is an important forage and potential bioenergy crop in Canada and world-wide. The development of heat stress-tolerant cultivars would be

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beneficial to reduce production loss as mean annual temperatures in Canada rise. Members of the highly conserved miR156 family regulate the Squamosa Promoter-Binding Protein-like (SPL) genes that are involved in plant growth and development and are differentially expressed in response to various stresses, including heat stress. In this study, we investigate the role of miR156 in heat stress tolerance using alfalfa overexpressing the miR156d precursor at 40°C. Overexpression plants showed reduced leaf relative water content and increased midday leaf water potential under heat stress after four days. Preliminary survival analysis data suggest miR156d overexpression may also increase heat stress tolerance. We will also present data on effect of heat on leaf chlorophyll and carotenoid contents, as well as expression analysis of miR156 and its seven target SPL genes in alfalfa subjected to heat stress over time. Specific miR156 targets differentially regulated under heat stress determined by ongoing expression analysis will be further investigated using overexpression and RNAi knockdown plants.

Craig Matthews [email protected]

105-AB: Physiological and epigenomic responses during and post abiotic stress in Populus balsamifera

Kayla Riane DIAS1*, Barb THOMAS2, Katharina BRÄUTIGAM 1,3 1. Department of Cell and Systems Biology, University of Toronto, Toronto, Ontario, Canada 2. Department of Renewable Resources, University of Alberta, Edmonton, AB, Canada 3. Department of Biology, University of Toronto Mississauga, Mississauga, Ontario, Canada The ability to respond efficiently to environmental change is vital to all forms of life, especially for long-lived plants such as trees. Here, we aim to understand how genotypes of a common Canadian tree, Populus balsamifera, respond to abiotic stress with emphasis on acute perturbation and stress recovery. We aim to integrate responses to two related stresses, at the physiological, molecular and epigenetic level. Based on preliminary screening for salt tolerance, three genotypes of the Alberta Pacific collection were selected and exposed to either osmotic or ionic stress for a period of two weeks, followed by one month recovery period for both stress treatments. Weekly assessment of growth and stomatal conductance indicated that all genotypes were severely stressed under strong osmotic stress conditions but were able to fully recover, while strong ionic stress resulted in a gradient of responses. These results illustrate that poplar genotypes can selectively exhibit a high degree of plasticity that allows them to acclimate to a changing environment. It also highlights the importance of understanding the mechanisms that facilitate tree survival, an information that might be utilized for informed planning in commercial plantations, forest ecosystems, or reclamation efforts. Kayla Dias ([email protected])

106-AB: Constitutive expression of an exogenous PHT1 family member in hybrid poplar Letitia DA ROS1*, Robert GUY2, Shawn D MANSFIELD1 1. Department of Wood Science, University of British Columbia, Vancouver, B.C. 2. Department of Forest and Conservation Sciences, University of British Columbia, Vancouver, B.C.

Leaf phosphate accumulation has been observed in different species of flowering plants, including barley (Hordeum vulgare), rice (Oryza sativa), and poplar (Populus spp.) when plants are exposed to high external concentrations. To maintain cellular phosphate homeostasis in excess conditions, plants are dependent on the storage capacity of the vacuole, and eventual net export of phosphate from plant roots to avoid phosphorus induced zinc and copper deficiencies. In plants adapted to phosphorus impoverished soils, phosphate accumulation occurs but they are largely unable to reduce phosphate uptake as internal concentrations increase. A continuing trend that demonstrated how plants adapted to low phosphorus conditions that moderate phosphate uptake show lower resorption efficiencies, suggesting constitutively expressed phosphate transporters may play dual roles in phosphate uptake from the soil and resorption from mature leaves.

From a phytoremediation perspective, should this relationship hold true for poplar species, it could be used to improve the phosphate storage capacity and resorption of trees growing on marginal soils with the intent of limiting phosphorus entry into riparian ecosystems. Transgenic hybrid poplar (P. alba x P. grandidentata),

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harbouring a constitutively expressed low affinity potato phosphate transporter (35S::StPT1) were generated using agrobacterium-mediated transformation. Of the transgenic lines created, only the highest expressing 35S::StPT1 line demonstrated a marked increase in phosphate accumulation when exposed to high external phosphate. These data suggest that phosphate mobilization from the vacuole during senescence in P39 is limited, thereby masking any potential effects of the plasma membrane bound phosphate transporter 1 (PHT1) family during remobilization. Letitia Da Ros [email protected] 107-AB: Effects of Heat Stress on Early Developmental Processes in Fucoid Algae Mary E. CLINTON1*, Sherryl R. Bisgrove1 * 1. Department of Biological Sciences, Simon Fraser University, Burnaby, BC

Fucoid algae are intertidal brown seaweeds of the class Phaeophyceae, found on coastlines worldwide. As canopy-forming macroalgae, they are foundation species, providing an essential microclimate in which many other marine organisms make their homes, find food and reproduce. Consequently, establishing an understanding of how they are being affected by rising global temperatures could provide insight into the effects of climate change on the intertidal community as a whole, with implications for conservation. Recently, sea surface temperatures (SSTs) in the Salish Sea have begun to rise, with some areas warming significantly more than others. The goal of this project is to examine the effects of warm temperatures on early development in fucoid algae. In order to do this, two main parameters will be assessed: the ability of zygotes to complete early developmental processes, and the production of reactive oxygen species (ROS) during development. Various stressors, including heat, have been found to induce oxidative stress in eukaryotic cells by stimulating overproduction of ROS. As such, measuring ROS production throughout early development will provide an indication of how heat stress is affecting the zygotes. By comparing early development and ROS levels between populations, we aim to assess whether Fucus populations growing at sites associated with warmer SSTs exhibit greater resilience to heat stress. If so, they would represent sources of high temperature tolerant stocks that may be useful in restoration efforts at sites where Fucus has been lost. Mary Clinton [email protected]

108-AB: Translating frost-tolerant seed degreening from Arabidopsis to Canola

Mendel Perkins1,2, Logan Skori1, Muhammad Jamshed1, Subramanian Sankaranarayanan1, Matija Stanic1*, Marcus Samuel1

1. Department of Biological Sciences, The University of Calgary, Calgary, AB 2. University of British Columbia, Department of Botany, Vancouver, BC

Non-lethal frost during critical stages of Brassica napus (Canola) embryo development is known to significantly increase seed chlorophyll levels. Increasing seed chlorophyll levels result in seed downgrading, which results in economic penalties to producers. Components of the seed de-greening pathway have been identified in the model organism Arabidopsis thaliana. ABI3, a transcription factor involved in mediating abscisic acid (ABA) responses has been shown to regulate a large suite of genes related to seed maturation and de-greening. During seed maturation, ABI3 is required for the transcriptional activation of SGR2, which encodes a magnesium dechelatase implicated in both chlorophyll and photosystem degradation. ABI3 has been shown to interact with the SGR2 promoter to drive transcriptional activity. In Arabidopsis, overexpression of ABI3 was sufficient to impart frost-tolerant seed degreening. Given the high level of sequence similarity of SGR2 and ABI3 between species, it is expected this technology can be translated into Canola. To accomplish this, Brassica napus homologs of SGR2 and ABI3 were isolated and tested for their ability to perform similar functions using appropriate Arabidopsis mutants. In parallel, transgenic Canola lines constitutively overexpressing ABI3 were developed. These transgenic lines can de-green more completely than wild-type lines following frost exposure. Despite being expressed ectopically, the downstream up-regulation of SGR2 was restricted to the seed suggesting additional regulation in the leaves prevents ABI3 overexpression from initiating premature leaf chlorophyll catabolism. In addition, estimations of the transgenic lines growth characteristics and yield suggest an absence of deleterious pleiotropic effects.

Matija Stanic [email protected]

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109-AB: Adaptations of Eelgrass to Living in Anoxic Environments Natasha H. FICZYCZ*1, Sherryl R. BISGROVE1, Nikki WRIGHT2 1. Department of Biological Sciences, Simon Fraser University, Burnaby, BC 2. SeaChange Marine Conservation Society, Victoria, BC Eelgrass (Zostera marina) beds create important habitats in many coastal estuaries in Canada. The species provides many ecosystem services for invertebrates and fish including refuge, food and shelter. Industrial, residential, and recreational activities have resulted in the loss of several eelgrass communities in local B.C. waters. In particular, forestry practices in British Columbia have created toxic environments in many areas that once housed thriving eelgrass communities. Marine sediments at sites with large amounts of wood waste are anoxic and high in H2S. This phytotoxin is produced by bacteria that use sulfate from the seawater as an electron acceptor during the breakdown of organic compounds. The SeaChange Marine Conservation Society is working to restore eelgrass at sites in B.C. from which it has been lost. This is done by transplanting shoots from native beds located near the restoration sites. We report here an assessment of eelgrass productivity at several of these restoration sites. Studies aimed at re-establishing eelgrass communities at sites impacted by high levels of wood waste are also underway. Eelgrass can adapt to growth in anoxic environments by developing large air spaces or lacunae in tissues called aerenchyma. These large air spaces increase oxygen diffusion from sites of photosynthesis in the leaves to the rhizomes buried in the sediment. This anatomical feature is being used to assess whether eelgrass growing near sites impacted by high levels of wood waste might be better adapted for growth in this environment than individuals from more pristine areas. We are determining whether there is a correlation between the size of lacunae in eelgrass aerenchyma and H2S levels in sediments, by sampling eelgrass sites that are as close as possible to contaminated areas as well as locations that are further away.

Natasha Ficzycz [email protected]

110-AB: Heterologous expression of Brassica napus miR395f gene in Arabidopsis thaliana affects cold tolerance

Swati Megha*, Urmila Basu, Nat N.V. Kav Department of Agricultural, Food and Nutritional Science, University of Alberta, Edmonton, AB T6G 2P5 Plants are exposed to a plethora of environmental stresses, including temperature extremes, which limit their growth, development and productivity. Nevertheless, plants have the plasticity in their defense mechanisms enabling them to tolerate and, sometimes, even survive adverse environmental conditions. MicroRNAs (miRNAs) are small non-coding RNAs, approximately 18-24 nt in length, and are being increasingly recognized as regulators of gene expression at the post-transcriptional level. They possess the ability to influence a broad range of biological processes including growth, development and stress responses in plants. Previously, next-generation sequencing of cold-stressed Brassica napus in our lab identified increased expression of miR395f. miR395f is predicted to target ATP sulfurylases (APS1, APS3 and APS4) which are ubiquitous enzymes catalyzing intracellular sulfate activation. qRT-PCR analysis revealed an increase in the expression of APS4 transcript and a decrease in the expression of APS1 and APS3 in cold stressed B. napus tissues. It is known from the literature that the over-expression of APS resulted in enrichment of glutathione – one of the most abundant antioxidants. Therefore, we hypothesized that an increase in miR395f will lead to an increased expression of APS4 and higher accumulation of glutathione, thereby imparting increased cold tolerance to plants. In the current study, we overexpressed the B. napus precursor-miR395f in Arabidopsis thaliana and investigated its effect on cold responses. Initial findings have revealed that overexpression of pre-BnamiR395f leads to lower survival rates and increased electrolyte leakage when compared to wild type plants. Work is in progress to further characterize the independent transgenic lines overexpressing pre-BnamiR395f in order to further investigate the molecular mechanisms underlying decreased cold tolerance in A. thaliana following the overexpression of this pre-miRNA.

Swati Megha [email protected]

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111-AB: Untangling salt stress on growth and photosynthetic performance of Salix spp. Xinyi HUANG1*, Raju SOOLANAYAKANAHALLY2, Robert GUY3, Shawn D. MANSFIELD1 1. Department of Wood Science, University of British Columbia, Vancouver, B.C. 2. Saskatoon Research and Development Centre, Agriculture and Agri-Food Canada, Saskatoon, S.K. 3. Department of Forest and Conservation Sciences, University of British Columbia, Vancouver, B.C. Soil health and sustainable soil management are vital determinants for plant productivity, and consequently societal well-being. High soil salinity and soil salinization are problems as ancient as the history of agricultural activities, and remain one of the major causes of soil degradation globally. As a consequence of escalating climatic conditions and the rapidly growing human population, food security and energy demand are significant global challenges, especially since saline soils often induce nutrient imbalance, specific ion toxicity and challenge osmotic adjustment of plants. On the Canadian prairies, soluble sulphate salts accumulate in the surface and subsurface soils. Willows (Salix spp.) have shown inherently high variation in salt tolerance and have been heavily involved in biomass production across Canada, and therefore offer a multipurpose feedstock for bioenergy applications grown on unproductive marginal lands. To examine the inherent variability, several native and hybrid willows were investigated for their tolerance levels to salt stress, by exploring the osmotic stress and sodium toxicity caused by saline soils. To this end, 12 hybrid and native willow genotypes were supplemented with four different salt (Na2SO4 and MgSO4) solutions for three months in perennial soil mix. Tree growth and photosynthetic responses were monitored during the growth trial, and tissue samples from different organs were harvested for elemental and metabolic analysis. It is clear that growth and photosynthesis were severely impacted by salt treatments, and furthermore, the performance of the willows depends on both concentration and the composition of the salt treatments. Xinyi Huang [email protected]

112-AB: A domesticated transposable element is essential for salinity defense in Arabidopsis thaliana Yang Shao, Thomas Bureau 1. Department of Biology, McGill University, Montreal, Quebec Canada H3A1B1 Transposable elements (TEs) were once thought to be junk DNA, with no benefits towards the host. They are classified into DNA transposons and retrotransposons. In recent decades some DNA transposon-derived genes were found to be functional in plants. These types of genes are defined as domesticated transposable elements (DTEs). The reported DTEs in plants including the MUSTANG and FAR1/FHY3 gene families, both of which evolved from the MULE TE superfamily, and the DAYSLEEPER gene, derived from the hAT TE superfamily, are transcriptional regulators and play important roles during plant development. Here we found MUG4, a member of the MUSTANG gene family, is essential for abiotic stress adaptation. Two T-DNA insertion lines were both shown to be hyper-sensitive to KCl and NaCl, but not sensitive to osmotic stresses induced by mannitol or PEG. Molecular complementation experiments confirmed the mug4 salt sensitive phenotype. These results suggest MUG4 may be specifically involved in ion toxicity defense but not directly involved in osmotic stress response. MUG4 was found to be in the nucleus, however, unlike FAR1 nor FHY3, MUG4 did not show transcriptional activation. All the results above indicate MUG4 has novel functions compared to previously reported DTEs, and opens a new door for valuable roles of DTEs in plant adaptation to stress. Yang Shao [email protected]

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113-AB: Expression of phytoglobin affects nitric oxide metabolism and energy state of barley plants exposed to anoxia Devin W. COCHRANE1, Kim H. HEBELSTRUP2, Abir U. IGAMBERDIEV1* 1. Department of Biology, Memorial University of Newfoundland, St. John's, NL, Canada 2. Department of Molecular Biology and Genetics, Aarhus University, Aarhus, Denmark Class 1 plant hemoglobins (phytoglobins) are upregulated during low oxygen stress and participate in metabolism and cell signaling via modulation of the levels of nitric oxide (NO). We studied the effects of overexpression and downregulation of the class 1 phytoglobin gene in barley (Hordeum vulgare L.) under low oxygen stress. The overexpression of phytoglobin reduced the amount of NO released, while downregulation significantly stimulated NO emission (measured by the chemiluminescent method). It has previously been shown that NO inhibits aconitase, so the decreased aconitase activity in downregulating plants can act as a biomarker for high internal NO levels. The overexpression of phytoglobin corresponded to higher ATP/ADP ratios, pyrophosphate levels and aconitase activity under anoxia, while the downregulation of phytoglobin resulted in the increased level of protein nitrosylation and led to elevation of alcohol dehydrogenase and nitrosoglutathione reductase activities. This proves that the hypoxically induced phytoglobin supports energy levels and prevents the signs of NO toxicity under the conditions of oxygen deficiency. The plants overexpressing phytoglobin were the only type to germinate and survive under hypoxia; however they showed various signs of stunted growth under the normoxic conditions. The results indicate that the upregulation of phytoglobin protects plant cells via NO scavenging and improves their low oxygen stress survival. However, phytoglobin overexpression may not be useful for cereal crop improvement since it comes with a significant trade-off of muddying normoxic NO signaling pathways. Abir Igamberdiev [email protected] 114-AB: Physiological basis of Seasonal Changes in Net Photosynthesis Rates, Transpiration rates, and Leaf Senescence in Catalpa speciose Aman CHERA1*, Jarnail CHANDI1, Santokh SINGH1 1. Department of Botany, University of British Columbia, Vancouver, B.C, Canada

The focus of this study was to examine the effects of environmental and developmental changes on photosynthesis and transpiration rates of Northern Catalpa (Catalpa speciose) leaves during late summer until fall season. In addition, we investigated the effects of various plant regulators, e.g. auxin (naphthaleneacetic acid or NAA), cytokinin (6-benzylamino purine or BAP), gibberellic acid (GA3), 2 chloro-ethyl-phosphonic acid (CEPA, an ethylene-releasing compound), abscisic acid (ABA), and jasmonic acid (JA) on leaf senescence in Northern Catalpa. As the season changed from summer to autumn the Northern Catalpa leaves started to show signs of leaf senescence. The net photosynthesis rates and transpiration rates of Northern Catalpa leaves progressively declined during the onset of autumnal senescence. NAA was the most effective plant regulator in delaying the senescence of the Northern Catalpa leaves. On the other hand, ABA was the most effective plant regulator in enhancing the senescence of the Northern Catalpa leaves. The physiological and biochemical basis of these research findings will be discussed. Aman Chera [email protected]

115-AB: The Effects of Soil Compaction on Photosynthesis, Transpiration and Leaf Senescence in Japanese Katsura (Cercidiphyllum japonicum) Amneet DHILLON1*, Jarnail CHANDI1, Santokh SINGH1 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada V6T 1Z4 This study focused on the effects of soil compaction, particularly in urban areas, on leaf senescence, net photosynthesis rates and transpiration rates in Cercidiphyllum japonicum leaves. The photosynthesis and

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transpiration rates were measured using the CI-340 Handheld Photosynthesis System. Leaf senescence was measured by analyzing relative chlorophyll levels using the CL-01 Chlorophyll Content System. The transpiration rates were relatively similar between the control and soil compaction treatment in the beginning of this study. The net photosynthesis rates and chlorophyll levels were generally lower in the trees experiencing soil compaction. The results of this study suggest that plants growing in soil compaction conditions may have experienced stress conditions including decreased access to essential nutrients like nitrogen. In summary, our study showed that the compaction of soil caused decreased photosynthesis rates and an overall yellowing of leaves of C. japonicum. Furthermore, this understanding can be applied towards using plants as a solution to the increasing carbon dioxide levels in the atmosphere. Amneet Dhillon [email protected]

116-AB: Betalains confer photoprotection in Amaranthus plants grown at high and low irradiance Elena BENIC1*, Karen K. TANINO1, Gordon R. GRAY1,2 1. Department of Plant Science, University of Saskatchewan, Saskatoon, SK 2. Department of Biochemistry, University of Saskatchewan, Saskatoon, SK Photoinhibition is defined as a light dependent decrease in photosynthetic efficiency and capacity. Plants have evolved numerous mechanisms to cope with photoinhibition and one of these photoprotective strategies involves the screening of incident light. The objective of this research was to evaluate the photoinhibitory responses of red and green vegetable varieties of Amaranthus spp. grown at high light (HL, 500 µmol m-2 s-1 photosynthetic photon flux density (PPFD)) and low light (LL, 70 µmol m-2 s-1 PPFD) and the role of pigmentation as a mechanism of tolerance to photoinhibition. Photosystem II photochemical efficiency (Fv/Fm) was measured for both HL and LL grown plants before and after exposure to a photoinhibitory irradiance (1400 µmol m-2 s-1) at 2°C for 4 hours. When grown at HL the red variety showed only a 20% loss of Fv/Fm while the green variety exhibited a 59% loss. Similarly, plant growth at LL resulted in 44 and 79% losses of Fv/Fm in the red and green varieties respectively. Upon the examination of photoprotective pigments at HL growth conditions, the red variety possessed a 2.6- and 4.7-fold higher content of carotenoids and betalains respectively in comparison to the green variety. A similar trend was observed for growth at LL with a 2.2-fold increase in carotenoids and a 3.5-fold increase in betalains evident in the red variety. These results demonstrate that in Amaranthus, HL growth promotes the accumulation of photoprotective pigments which in turn increase tolerance to photoinhibition. Elena Benic [email protected] 117-AB: Extraorgan freezing in cold acclimated winter wheat and rye: Role of the leaf sheath in crown freezing survival Ian R. WILLICK1, D. Brian FOWLER1, Karen K. TANINO1 1. Department of Plant Sciences, University of Saskatchewan, Saskatoon, SK The most critical region for winter cereal winter survival is the crown. Freezing results in the disruption of crown structure as apoplastic ice grows at the expense of intracellular water. Cold acclimated crowns are more resistant to freezing due to less free water in the vascular transition zone (VTZ) and shoot apical meristem (SAM) meristematic regions, reducing the formation of ice and subsequent amount of physical damage. Remarkably, little work has been done to investigate the role of the leaf sheath that surrounds the crown, and how its freezing affects overall crown injury. We hypothesize that in cold acclimated crowns, the leaf sheath acts as an ice sink to mitigate freezing injury to the VTZ and SAM. Patterns of ice accumulation and injury were examined using ice sectioning, SEM and tetrazolium chloride vital staining in winter wheat (Triticum aestivum L. cv ‘Norstar’) and rye (Secale cereale L. cv ‘Puma’). By manipulating the crown, we identified that the absence of the leaf sheath resulted in damage to the SAM and reduced recovery in cold acclimated ‘Norstar’ but not ‘Puma’. These results were validated with tissue specific freezing tests, differential thermal analysis and ice activity assays. It is postulated that leaf sheath ice nucleation activity levels contribute to the establishment of whole crown freezing behaviours by ensuring the order

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of freezing among the tissues, from the primary freeze in non-critical regions to the final freezing event in critical tissues. Ian R. Willick [email protected]

118-AB: Heat stress affects seed development by modulating gibberellin biosynthesis and metabolism pathways in pea (Pisum sativum L.)

Harleen KAUR, Jocelyn A. OZGA, Dennis M. REINECKE Plant BioSystems Group, Department of Agricultural, Food, and Nutritional Science, University of Alberta, Edmonton, AB, Canada T6G 2P5

Pea (Pisum sativum L.) is an important crop grown in the western Canadian prairies where summer temperatures during reproductive development can adversely affect fruit/seed yield. The developing floral buds and open flowers borne on first 5 flowering nodes of pea plants (cv. Carneval) exposed to moderate heat stress (35oC for 6 h) for 4 days during early flowering exhibited increased % seed/ovule abortion producing more pods with reduced length and fewer seeds (2-4 seeds/pod) at maturity than plants grown under normal temperature (19oC) conditions (5-6 seeds/pod). The number of seeds within the fruit is tightly linked to fruit/pod size in pea. Fruit and seed development are complex processes that are regulated by plant hormones including gibberellins (GAs). We further explored if part of the inhibitory effect of high temperature on fruit/seed growth is through modulation of GA biosynthesis. Bioactive GA levels were substantially more abundant in the developing seeds than other fruit tissues. GA profiling suggested that when the plants were exposed to the higher temperature, the production of early 13-hydroxylated bioactive GA precursors (GA19 and GA20) was reduced and inactivation of bioactive GA1 increased leading to lower bioactive GA1 and GA3 levels in the seeds. Bioactive non-13-hydroxylated GAs, GA4 and GA7 were also lower in seeds when the plants were exposed to heat stress during early flowering. These data support that an increase in the % seed abortion induced by high temperature is associated with reduced GA biosynthesis and increased bioactive GA inactivation capacity in the developing seeds.

Presenter: Jocelyn Ozga [email protected]

119-AB: Investigation of physical and biochemical changes in the adaxial epicuticular layer of mature leaf following chilling pre-treatment in Zea mays

Kaila J. HAMILTON1, Karen K. TANINO1 1. Department of Plant Sciences, College of Agriculture and Bioresources, University of Saskatchewan, Saskatoon, Saskatchewan, Canada Zea mays is considered sensitive to chilling and freezing stress. Yield-reducing frost damage in corn has limited expanded production into regions such as the Canadian prairies. The leaf epicuticular layer can serve as a barrier for avoidance of ice nucleation and inhibition of ice propagation but the influence of the preceding fall temperatures on epicuticular properties and ice nucleation have not been previously investigated. Using Attenuated Total Internal Reflectance Fourier Transform Spectroscopy (ATR-FTIR), the lipid composition of mature grain corn leaves was studied under controlled-environment and field conditions to evaluate biochemical changes induced by chilling pre-treatment in the epicuticular layer. Plants were exposed to chilling pre-treatment conditions based on the 30-year average (18°C/6°C) over a 10 day period preceding frost. Temperature of ice nucleation was determined using thermal imaging. We are developing a model using complementary techniques using physical trait inputs as predictors to estimate temperature of ice nucleation. The hydrophobic properties of the leaf surface are determined by the biochemical makeup of the epicuticular layer. Hydrophobicity has been linked to increased freezing avoidance. ATR-FTIR results showed changes following chilling pre-treatment in the asymmetrical stretching region of the CH2 group associated with aliphatics which comprise the plant cuticle (cutin, waxes and cutan). A reduction in leaf surface hydrophobicity was observed following chilling pre-treatment. Zea mays represents a useful model to evaluate the impact of abiotic stresses, specifically the unstudied impact of chilling pre-treatment on epicuticular wax modification and subsequent ability to avoid freezing using a whole plant system.

Kaila Hamilton [email protected]

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120-AB: Preharvest treatment application: A new era for UV-C hormesis Marie Thérèse CHARLES1, Yanqun XU 1,2, Zhichun XIE1,3, Joseph ARUL4, Zisheng LUO2. 1. Saint-Jean-sur-Richelieu Research and Development Centre, Agriculture and Agri-Food Canada, Saint-Jean-sur-Richelieu, QC, J3B 3E6, Canada 2. College of Biosystems Engineering and Food Science, Zhejiang University, Hangzhou 310058, People’s Republic of China 3. College of Forestry, Northwest A&F University, Yangling, Shaanxi 712100, China 4. Horticultural Research Center and Department of Food Science and Nutrition, Université Laval, Sainte-Foy, Québec, Canada G1K 7P4 Hormesis is the phenomenon that underlies a set of favorable responses observed in a biological system subjected to low levels of exposure to stressors. Over the last thirty years, hormetic levels of short-wave ultraviolet (UV-C) radiations, applied at the post-harvest stage, have been studied for their potential to extend shelf life and improve quality of a wide range of horticultural products. UV-C hormesis has been found to be effective in delaying senescence, inactivating pathogenic and deterioration causing microorganisms, preventing losses due to rot and disease development, and improving the bioactive components of the products. Unfortunately, in the case of fragile commodities, the non-systemic action of UV-C slows down the adoption of this practice by the fresh fruit and vegetable (F & V) industry. Recent considerations have been given to F & V responses to UV-C hormesis when applied at the prehavest stage. Early findings highlighted that this new approach could be considered in pest management and as a sustainable way to improve fresh F & V quality and composition. [email protected]

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Biotic Stress Posters (121-‐BS to 141-‐BS) Wednesday July 5, FSC Atrium 11:20am -12:30pm

121-BS: Potential role of narigenin chalcone in plant disease resistance

Igor D. ALBUQUERQUE1, Yangdou WEI1, Kenneth WILSON1.

1. Department of Biology, University of Saskatchewan, Saskatoon SK

During studies of Arabidopsis mutants defective in the phenylpropanoid pathway, the tt5 mutant (chalcone isomerase) was found to have reduced growth, but enhanced resistance to Powdery mildew (Erysiphe cichoracearum) and Anthracnose (Colletotrichum higginsianum). Chalcone isomerase is a key enzyme in flavonoid biosynthesis. Interestingly, if the previous gene (chalcone synthase - tt4) is knocked out no flavonoids are produced, but the plants do not exhibit enhanced disease resistance. The double mutant (tt4/tt5) also has normal disease susceptibility. We hypothesize that a chemical product differentially accumulated by the tt5 plants is responsible for its phenotype. Based on current knowledge of flavonoid biosynthesis this compound should be narigenin chalcone. However, during initial experiments we found that tt5 plants exhibited cell death patterns consistent with salicylic acid accumulation. Salicylic acid is a hormone associated with disease resistance through activation of the plant immune system. The next steps of my project are: quantify flavonoid components in tt5 leaf extracts using high performance liquid chromatography, and examine the expression of salicylic acid, and other disease inducible genes using RNAseq.

Igor de Albuquerque [email protected]

122-BS: Microarray analysis of lodgepole and jack pine seedlings responding to inoculation by mountain pine beetle fungal associate Grosmannia clavigera under well watered and water deficit conditions. Elizabeth MAHON1*, Janice COOKE2, Adriana ARANGO-VELEZ3 1. Department of Wood Science, University of British Columbia, Vancouver, B.C. 2. Department of Biological Sciences, University of Alberta, AB, Canada 3. Department of Forestry & Horticulture, The Connecticut Agricultural Experiment Station, CT To date mountain pine beetle (MPB) has affected more than 28 million ha. of pine forests in western North America. Lodgepole pine (Pinus contorta) has been the primary species affected by the current outbreak, however as MPB range expands eastward beyond its historical habitat, the bark beetle has encountered a novel host: jack pine (Pinus banksiana). Ecological evidence has indicated that host trees originating from MPB’s historic range have lower host quality compared to hosts from novel habitats, suggesting that co-evolved lodgepole pine may have acquired induced and constitutive defenses against MPB that are not present in jack pine. Additionally, ecological evidence suggested that trees subjected to abiotic stresses such as drought are more susceptible to MPB attack. Using data generated from a large-scale microarray study, we examined the transcriptomic response of both lodgepole and jack pine seedlings inoculated with MPB fungal associate G. clavigera in order to investigate the genetic mechanisms underlying these ecological trends. In both species, activation of defense response pathways occurred through signaling action of jasmonic acid and ethylene. We identified qualitative differences between the secondary metabolite biosynthesis genes induced by lodgepole and jack pine in response to G. clavigera, with lodgepole pine seedlings exhibiting induction of more genes involved in flavonoid biosynthesis, and jack pine seedlings exhibiting induction of more genes involved in terpene biosynthesis. Our results also revealed that water deficit substantially impacts the transcriptomic response of both species to G. clavigera, however water deficit impacts lodgepole to a greater extent than jack pine. Elizabeth Mahon [email protected]

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123-BS: Natural variation of disease resistance to the Pseudomonas syringae effector HopX1 in Arabidopsis thaliana Natalie HOFFMANN1*, Bradley LAFLAMME1, Darrell DESVEAUX1,2 1. Department of Cell and Systems Biology, University of Toronto 2. Centre for the Analysis of Genome Evolution and Function, University of Toronto Pseudomonas syringae is a Gram-negative bacterial pathogen that causes disease in many plant species, including the model plant Arabidopsis thaliana. P. syringae is able to directly inject virulence proteins called effectors into plant cells through a type III secretion system to suppress plant immunity and cause disease. Plant species have evolved to recognize effectors using a diverse array of Resistance (R-) genes, which activate a robust immune response. HopX1, a member of the HopX family of effectors, is a cysteine protease that targets the jasmonic acid plant hormone pathway and suppresses immunity in susceptible plants. However, mechanisms of HopX1 detection in resistant plants remain largely uncharacterized. In order to better define the genetic and molecular basis of HopX1 recognition, we cataloged the natural variation of HopX1 resistance in an infection assay of global A. thaliana accessions collected by the 1001 Genomes Project. We hypothesized R-gene diversity between accessions would lead to altered disease susceptibility to infection with P. syringae pv. tomato DC3000 expressing HopX1 (PtoDC3000HopX1). In an infection assay of 86 accessions, we identified 17 accessions (20%) with enhanced resistance to PtoDC3000, and two accessions, Stepn-2 and Vie-0, with increased specific resistance to PtoDC3000HopX1. Resistant accessions showed significantly lower leaf chlorosis following infection compared to the Columbia-0 control. Future studies with Stepn-2 and Vie-0 could identify genetic polymorphisms underlying HopX1 recognition and give insight into disease resistance in A. thaliana. Natalie Hoffmann [email protected]

124-BS: The mitochondrial tail-anchored proteins, AtTTM1 and 2, are involved in senescence and immunity-related programmed cell death

Purva Karia1*, Huoi Ung1, Wolfgang Moeder1, Kazuo Ebine3, Anissa Poleatewich2, Michael Pautler2, Travis Banks2, Daryl Somers2, Takashi Ueda3, and Keiko Yoshioka1

1. Department of Cell &Systems Biology, University of Toronto, Toronto, Ontario 2. Vineland Research and Innovation Centre, Vineland Station, ON, Canada 3. National Institute for Basic Biology. Nishigonaka 38, Myodaiji, Okazaki 444-8585 Aichi, Japan

Triphosphate tunnel metalloenzymes (TTMs) comprise a superfamily of enzymes that hydrolyze organophosphate substrates. The members of this superfamily are found in all domains of life and the Arabidopsis thaliana genome encodes three TTM genes (AtTTM1-3). Unlike other members of this family, two of these isoforms, AtTTM1 and 2, hydrolyze pyrophosphate in vitro, making them the only TTMs characterized so far to possess pyrophosphatase activity. Confocal microscopy analysis unveiled their sub-cellular localization at the mitochondrial outer membrane. However, despite their same enzymatic activity and sub-cellular localization, knockout mutant analyses revealed different biological functions. AtTTM2 plays a role in pathogen resistance whereas AtTTM1 is involved in leaf senescence. Knockout mutants of AtTTM1 exhibit delayed dark-, ABA-induced and natural senescence whereas transient overexpression of AtTTM1 caused senescence-like cell death upon dark treatment indicating the involvement of AtTTM1 in the senescence pathway as a positive regulator. The ttm1 ttm2 double mutant displayed the same degree of delayed senescence and enhanced pathogen resistance as their respective single mutants, further confirming their roles in distinct biological processes. Together, these data suggest a novel connection between immunity-related programmed cell death and senescence through these mitochondrial membrane-localized tail-anchored proteins. To explore their agricultural application, EMS-mutagenized populations of several crop plants were created and mutants in two TTM2 paralogs in tomato and pepper were identified through DNA melting for genotyping and Illumina-based sequencing. A tomato ttm2 mutant displayed increased resistance to powdery mildew infection indicating the conserved function of TTM2 among crops.

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Keywords: Mitochondria, Tail-anchored protein, Triphosphate tunnel metalloenzymes (TTMs), programmed cell death, senescence, pathogen

Name: Purva Karia Email: [email protected]

125-BS: Plant Volatiles in Insect Pest Control: A promising new tool for IPM Scott HUGHES1,2*, Ian SCOTT1, Mana CROFT1, Tim McDOWELL1, Mark BERNARDS2, Abdelali HANNOUFA1 1. London Research and Development Centre, Agriculture and Agri-Food Canada, London, Ontario, Canada 2. University of Western Ontario, Department of Biology, London, Ontario, Canada. Integrated pest management (IPM) aims to use multiple control mechanisms in order to reduce insect pest populations in a sustainable fashion. The utilization of transgenic plants as a potential new tool for IPM is a growing area of interest. Plant volatile organic compounds (VOCs) play an important role in plant-insect interactions, and these interactions can be influenced and exploited by manipulating the expression of the genes involved. Previously, our group showed that overexpressing carotenoid cleavage dioxygenase (CCD) family members increases the production of repellent apocarotenoid volatiles in Arabidopsis thaliana. As a continuation of this work, we have successfully produced tomato Solanum lycopersicum plants overexpressing AtCCD4. Expression analysis of AtCCD4-OE lines showed a wide range of overexpression levels. Static headspace sampling of leaf VOC emissions using solid phase micro-extraction (SPME) and subsequent gas chromatography-mass spectrometry (GC-MS) revealed differences in VOC profiles between empty vector (EV) control and AtCCD4-OE plants. In initial dual-choice oviposition bioassays using greenhouse whitefly Trialeurodes vaporariorum Westwood (Hemiptera: Aleyrodidae) as a generalist pest model, AtCCD4-OE plants showed a trend, albeit statistically insignificant, towards having a repellent effect. Future work will focus on VOC analysis of tomato flowers and fruits, additional dual-choice and no-choice oviposition bioassays, and Y-tube olfactometer assays. Long term, plants with altered VOC profiles could act as trap crops or pest repellent crops within a “push-pull” IPM strategy, and could additionally aid in the maintenance of beneficial insect populations within a crop. Scott Hughes [email protected] 126-BS: Structure-function analysis of plant immune regulators SNC2 and BDA1

Weijie HUANG1, Yuli DING1 and Yuelin ZHANG1

1. Department of Botany, University of British Columbia, Vancouver, BC, Canada

Arabidopsis SNC2 (for Suppressor of NPR1, Constitutive2) encodes a receptor-like protein with important roles in plant immunity. A gain-of-function mutation in SNC2 leads to constitutive activation of defense responses in snc2-1D mutant plants. BDA1 (for bian da; “becoming big” in Chinese) encodes a protein with amino-terminal ankyrin repeats and carboxyl terminal transmembrane domains that is required for SNC2-mediated immunity. From snc2-1D suppressor screens, we identified 32 mutant alleles of SNC2 and 65 mutant alleles of BDA1 that suppress the autoimmune phenotypes in snc2-1D. Structure-function analysis of SNC2 and BDA1 is being carried out using these mutants and the results will be presented.

Weijie Huang [email protected]

127-BS: Perception of salicylic acid in the non-vascular plant Physcomitrella patens Yujun PENG1, Yuellin ZHANG1 1. Department of Botany, University of British Columbia, BC Salicylic acid (SA) is a signal molecule that induces defense responses against microbial pathogens in higher plants. In Arabidopsis, two groups of SA receptors, NPR1 and NPR3/NPR4, are involved in the perception of SA. In the

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lower plant Physcomitrella patens, SA treatment also induces defense gene expression. How SA is perceived in Physcomitrella patens is unknown. Blast analysis revealed that there is only one gene encoding a protein with high similarity to NPR1 and NPR3/NPR4 in the genome sequence of Physcomitrella patens. To test whether this NPR-like gene encodes a protein with similar function as NPR1 or NPR3/NPR4, we generated a construct expressing the gene under the 35S promoter and transformed it into npr1 and npr3 npr4 mutants to determine whether it can complement the mutant phenotypes of npr1 or npr3 npr4. Results from the complementation analysis will be reported. Our study may shed new light on how SA is perceived in non-vascular plants. [email protected]

128-BS: Characterization of a pipecolic acid biosynthesis pathway required for systemic acquired resistance

Pingtao DING1,6, Dmitrij REKHTER2,6, Yuli DING1,6*, Kirstin FEUSSNER2, Lucas BUSTA5, Sven HAROTH2, Shaohua XU3, Xin LI1, Reinhard JETTER1,5, Ivo FEUSSNER 2,4,*, Yuelin ZHANG1,* 1 Department of Botany, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 2 Georg-August-University, Albrecht-von-Haller-Institute for Plant Sciences, Department of Plant Biochemistry, D-37073 Goettingen, Germany 3 National Institute of Biological Sciences, Beijing, 102206, China 4 Georg-August-University, Goettingen Center for Molecular Biosciences (GZMB), Department of Plant Biochemistry, D-37073 Goettingen, Germany 5 Department of Chemistry, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 6 These authors contributed equally to this work. Systemic acquired resistance (SAR) is an immune response induced in the distal parts of plants following defense activation in local tissue. Pipecolic acid (Pip) accumulation orchestrates SAR and local resistance responses. Here we report the identification and characterization of SAR-Deficient 4 (SARD4), which encodes a critical enzyme for Pip biosynthesis in Arabidopsis. Loss of function of SARD4 leads to reduced Pip levels and accumulation of a Pip precursor, Δ1-piperideine-2-carboxylic acid (P2C). In E. coli, expression of the aminotransferase ALD1 leads to production of P2C and addition of SARD4 results in Pip production, suggesting that a Pip biosynthesis pathway can be reconstituted in bacteria by co-expression of ALD1 and SARD4. In vitro experiments showed that ALD1 can use L-lysine as a substrate to produce P2C and P2C is converted to Pip by SARD4. Analysis of sard4 mutant plants showed that SARD4 is required for SAR as well as enhanced pathogen resistance conditioned by overexpression of the SAR regulator FMO1. Compared to wild type, pathogen-induced Pip accumulation is only modestly reduced in the local tissue of sard4 mutant plants, but it is below detection in distal leaves, suggesting that Pip is synthesized in systemic tissue by SARD4-mediated reduction of P2C and biosynthesis of Pip in systemic tissue contributes to SAR establishment. Yuli Ding [email protected]

129-BS: Understanding how Pectin Modifying proteins BGAL4 and BXL4 contribute to defence against Pseudomonas syringae and Hyaloperonospora arabidopsidis in Arabidopsis Athanas GUZHA1, Marcel WEIMER2, George HAUGHN3, Till ISCHEBECK1 1. Department of Plant Biochemistry, Albrecht-von-Haller-Institute for Plant Sciences, Georg-August-University, Goettingen, Germany 2. Department of Plant Cell Biology, Schwann-Schleiden Centre, Georg-August-University, Goettingen, Germany 3. Department of Botany, University of British Columbia, Vancouver, BC, Canada Pectin constitutes a major component of the primary cell wall in most dicotyledonous plant species. This pectin forms a barrier that plant pathogens have to breach in order to access the plant cell contents. Plants secrete pectin to the apoplast together with some cell wall proteins, including pectin modifying enzymes. Work on the pectin rich

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mucilage of Arabidopsis seeds has shown that these modifying enzymes are important for mucilage production with the right chemical composition and function as shown for BXL1 and MUM2 (of the BGAL family). Two pectin modifying genes xylan 1,4-β-xylosidase (Bxl4) and β-galactosidase 4 (Bgal4), which are members of Arabidopsis GH3 and GH35 families respectively show an upregulation of expression upon Pseudomonas syringae infection. Arabidopsis mutant lines deficient in either genes showed a significant increase in infection to Psuedomonas syringae and Hyapelonospora arabidopsidis compared to the wildtype. The observed heightened susceptibility points to their possible role in re-modeling of the cell walls in response to attack from the two pathogens. Whilst various roles of these genes in cell wall modification during plant development have been articulated, the precise role played by the genes is unknown, especially with regard to immune responses to pathogen attack. This ongoing research intends to elucidate the molecular mechanisms underlying this observed interaction. Athanas Guzha [email protected]

130-BS: Foliar selenium application reduces late blight severity and incidence and induces the phenolic pools in potato leaves and tubers

Ashok SOMALRAJU1,2, Jason MCCALLUM1, David MAIN1, Rick PETERS1, Bourlaye FOFANA1

1. Charlottetown Research and Development Centre, Agriculture and Agri-Food Canada, 440 University Avenue, Charlottetown, Prince Edward Island, C1A 4N6, Canada

2. University of Prince Edward Island, 550 University Avenue, Charlottetown, Prince Edward Island, C1A 4P3, Canada

Potato is the 4th most consumed food crop worldwide and is central to global food security. However, potato production faces many constraints including poor yield and quality, diseases, and pest pressure. Among potato diseases, late blight is one of the most devastating pathogens causing a serious threat to potato production. Whereas disease control by fungicides appears to be effective in conventional production systems, organic systems are still looking for viable disease control options. Selenium is a mineral micronutrient essential to humans and animals in trace amounts, and is widely used for crop and livestock biofortification. Nonetheless, its essential role for plant growth is not yet well-established, despite its role as antioxidant, in delayed senescence and protection against fungal infection and aphid feeding. As an alternative to chemical fungicides, pesticides derived from certain minerals such as silicon and selenium may be options for organic production. To date however, the role of selenium on potato late blight is unknown. This study was designed to evaluate the effect of selenium on late blight in potato plants grown in greenhouse, and to determine some of its modes of action in the plants. Our preliminary results show that foliar selenium applications reduce the severity and incidence of late blight and interfere with the phenylpropanoid pathway, inducing the production of secondary metabolites including anthocyanins and hydroxyl-cinnamic acids in the leaves and tubers of treated plants compared to non-treated plants. These results suggest that selenium may contribute to improved potato late blight control.

Bourlaye Fofana [email protected]

131-BS: A Truncated NLR Protein Is a MOS6/IMPORTIN-α3 Interaction Partner and Required for Plant Immunity Daniel LUEDKE1, Charlotte ROTH1, Melanie KLENKE1, Annalena QUATHAMER1, Oliver VALERIUS2, Gerhard H. BRAUS2 and Marcel WIERMER1 1. Georg-August-University Goettingen, Albrecht-von-Haller Institute for Plant Sciences, RG Molecular Biology of Plant-Microbe Interactions, Julia-Lermontowa-Weg 3, 37077 Goettingen, Germany 2. Georg-August-University Goettingen, Institute for Microbiology and Genetics, Department of Molecular Microbiology and Genetics, Griesebachstrasse 8, 37077 Goettingen, Germany Importin-α proteins mediate the translocation of nuclear localization signal (NLS)-containing proteins from the cytoplasm into the nucleus through nuclear pore complexes (NPCs). Genetically, Arabidopsis IMPORTIN-α3/MOS6 (MODIFIER OF SNC1, 6) is required for basal plant immunity and constitutive disease resistance activated in the auto-immune mutant snc1 (suppressor of npr1-1, constitutive 1), suggesting that MOS6 plays a role in nuclear

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import of proteins involved in plant defense signaling. To identify potential defense-regulatory cargo proteins and interaction partners of MOS6, in silico database analyses and affinity-purification of functional epitope-tagged MOS6 from stable transgenic plants coupled with mass spectrometry (MS) have been conducted. Among thirteen candidate MOS6 interactors that were selected for functional analyses, a TIR-NBS (TN)-type protein is required for resistance against Pseudomonas syringae pv. tomato (Pst) DC3000 lacking the type III effector proteins AvrPto and AvrPtoB. When expressed transiently in N. benthamiana leaves, the TN protein binds to MOS6 but not to its closest homolog IMPORTIN-α6, and localizes to the endoplasmic reticulum (ER), consistent with a predicted N-terminal transmembrane domain in the TN protein. Our work uncovered a TN type protein as a new component of plant innate immunity that selectively binds to MOS6/IMPORTIN-α3 in planta. We speculate that release of the TN protein from the ER-membrane upon pathogen challenge and its subsequent nuclear translocation is important for maintenance of basal resistance to Pst DC3000 (ΔAvrPto/AvrPtoB). Daniel Luedke [email protected] 132-BS: Comprehensive phenotypic analysis of sobir7-1 mutant reveals an important regulatory role of the Carboxyl-terminus (CT) extension of BRI1-ASSOCIATED RECEPTOR KINASE 1 (BAK1) Di Wu*, Fan Xu, Yuelin Zhang Department of Botany, University of British Columbia, Vancouver, BC, Canada BAK1 (BRI1-ASSOCIATED RECEPTOR KINASE 1) plays essential roles in various developmental and immune signaling pathways. It harbors a carboxyl terminus (CT) extension outside of the kinase domain. The importance of this CT region is unknown in the context of plant growth and immunity. Here we report the comprehensive phenotypic characterization of sobir7-1, a novel BAK1 mutant allele, which causes deletion of the CT region. Our data demonstrate that BAK1 CT is required for FLS2- and EFR-mediated PAMP-triggered immunity (PTI), but is dispensable in BAK1/BKK1-inhibited cell death signaling and most likely, Brassinosteroid-mediated developmental processes. Regarding the mechanisms underlying the regulatory role of BAK1 CT region, we also showed that PTI defects caused by sobir7-1 mutation are not due to compromised protein stability, or impaired physical interaction with BOTRYTIS-INDUCED KINASE1 (BIK1). Furthermore, a survey of all putative LRR-RLKs in A. thaliana, revealed that a large number of LRR-RLKs contain carboxyl terminal region outside of the kinase domain, suggesting that the CT extension may play a general role in LRR-RLK-mediated signaling. Di Wu [email protected] 133-BS: Monaghan Lab: Plant immunology and immune homeostasis Jacqueline Monaghan Department of Biology, Queen’s University, Kingston ON Plants have evolved a multi-faceted immune system to fight against pathogen infection. While necessary for survival, pathogen perception and the activation of immune responses are energetically taxing for the host and have been linked to considerable fitness costs. Although defense signaling pathways must therefore be tightly regulated, very little is known about the biochemical mechanisms that tailor signaling to maintain cellular homeostasis. Our new research program at Queen’s University focuses on understanding the basic mechanisms that allow plants to defend against a vast array of potential pathogens while maintaining normal growth and development. To this end, our work address the following biological questions using varied approaches: (1) What is the role of and interplay between different post-translational modifications on proteins involved in immune homeostasis? (2) What are the key regulators maintaining immune homeostasis and how do they function biochemically at the molecular level? (3) What developmental pathways are affected by immune signaling? Understanding the complexity of signaling events that underlie immune systems is integral to combating plant diseases that threateb food security world-wide. Contact: [email protected]

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134-BS: Linking genetic information to Colorado potato beetle resistance related metabolites in potato Jamuna R. PAUDEL1, Kyle GARDNER1, Benoit BIZIMUNGU1, Agnes MURPHY1, David D. KOEYER1, Jun SONG2, and Helen H. TAI1 1. Fredericton Research and Development Centre, Agriculture and Agri-Food Canada, New Brunswick 2. Kentville Research and Development Centre, Agriculture and Agri-Food Canada, Nova Scotia Colorado potato beetle, Leptinotarsa decemlineata (Say), is an economically important pest of cultivated potato. The adult and larvae CPB can defoliate entire field and significantly affect tuber yield. Chemical pesticides are commonly used to control CPB but insecticide resistant populations are increasing rapidly. Steroidal glycoalkaloids (SGAs) are important secondary metabolites in potato that have antimicrobial, antifungal and insect deterrent properties, and, therefore, these metabolites can be developed as an alternative method of pest management. However, due to SGA toxicity to humans at higher levels, reducing SGA levels in potato tubers is one of the important breeding targets. Genetic and environmental factors affect SGA content in potato; α-solanine and α-chaconine are the most abundant SGAs in cultivated potato, while diverse kinds of SGAs exist in wild relatives. Apart from abundance, the nature and composition of SGAs in potato foliage can affect CPB. Wild Solanum species, S. chacoense and S. oplocense contain Leptines and dehydrocommersonine, respectively that is responsible to provide host resistance. It has been found that the ratio of α-solanine and dehydrocommersonine is important for CPB resistance. Thus, these compounds are metabolic phenotype for CPB resistance. We have generated potato lines by crossing cultivated potato (S. tuberosum cv shepody) with F1 hybrid having S. oplocense background. Foliar metabolic profiling showed diverse levels of total glycoalkaloids and varying levels of α-solanine, α-chaconine and dehydrocommersonine in the backcross population. By incorporating these phenotypic information, bulk segregant analysis using next generation sequencing will be done to explore genomic regions in potato responsible for CPB resistance.

Jamuna Risal Paudel [email protected]

135-BS: Requirement for translationally regulated candidate genes during plant NB-LRR- mediated defense responses

Peter Moffett1 Teura Barff1, Mathias Cohen1, Mohamed El Oirdi1, Keiko Yoshioka2, and Louis-Valentin Meteigner1. 1Département de Biologie, Université de Sherbrooke, 2500 Boulevard de l’université, Sherbrooke, Québec, Canada, J1K 2R1 2University of Toronto, Department of Cell and Systems Biology, 25 Willcocks St. Toronto, Canada

Plants possess multiple defense mechanism, including the recognition of pathogen-encoded effectors by NB-LRR proteins, which induces Effector-Triggered Immunity (ETI). Much remains to be understood about downstream signaling in ETI, although we have previously shown that NB-LRR activation results in the translational repression of viral RNAs. To determine if translational control may also affect endogenous mRNAs we analysed the translatome upon activation of the NB-LRR protein, RPM1 using Translating Ribosome Affinity Purification followed by RNAseq (TRAPseq) in Arabidopsis thaliana. We find that the translation signature during an ETI response reflects a global shift away from growth and basic metabolism towards an upregulation of defense responses. Interestingly, several transcripts encoding proteins putatively involved in the Target of Rapamycin (TOR) pathway, a central regulator of protein translation, are regulated by this response. To determine the importance of translationally regulated genes in plant defense, we infected multiple A. thaliana knock out lines of selected candidate genes with Pseudomonas syringae pv tomato (Pto) and Hyaloperonospora arabidopsidis (Hpa). Using these assays, we found that many candidate genes are required for basal and/or NB-LRR mediated immunity. Indeed, suppression of TOR, genetically or pharmacologically, leads to increased resistance to pathogens while overexpression of TOR results in increased susceptibility, indicating an important role for translational control in the growth to defense switch. These studies have allowed us to identify new actors in plant defense pathways and future studies will focus on how their translation is regulated by NB-LRR signaling and how their protein products contribute to plant defenses.

Peter Moffett, [email protected]

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136-BS: Changes in wheat rachis composition after Fusarium infection in a resistant and a susceptible variety S. Shea MILLER, Mehri HADINEZHAD Ottawa Research and Development Centre, Agriculture & Agri-Food Canada, Ottawa, ON K1A 0C6 Fusarium head blight (FHB) is a fungal disease which affects both yield and quality of wheat. We are investigating changes in rachis composition after inoculation with Fusarium graminearum in a susceptible wheat, Chinese Spring (CS), and an addition line, CS-7EL, which contains a chromosome fragment from Thinopyrum elongatum carrying FHB resistance. Florets were point inoculated at anthesis; differences in severity of infection were apparent by day 5. Heads were collected at day 7, the florets were removed, and the rachises were sequentially extracted to give hot water soluble (free sugar), lignin, and holocellulose fractions. Gravimetrically, the largest fraction in all samples was holocellulose (56.8-64.6%), followed by hot water solubles (19.7-24.0%) and lignin (15.7-19.7%). In Fusarium inoculated samples, the hot water fraction increased in CS, suggesting cell wall degradation, while in CS-7EL, the lignin fraction increased, suggesting cell wall fortification. The hot water and holocellulose fractions were evaluated by HPLC-ELSD. Sucrose, glucose and fructose were the main sugars in the hot water fractions. After infection, glucose and fructose increased in both varieties: 70% and 87% respectively for CS, 45% and 72% respectively for CS-7EL. Hydrolysis of the holocellulose fractions showed the main cell wall polysaccharides in the rachis are cellulose and arabinoxylan. 7 days after infection, both glucose and xylose had decreased in CS, suggesting degradation of both cell wall polysaccharides; these sugars were unchanged CS-7EL, suggesting possible protection of the cell wall by increased lignification. Uronic acids in the holocellulose fraction increased in CS-7EL after infection, but decreased in CS. Shea Miller [email protected]

137-BS: Pathogen-induced cell wall remodeling and production of Danger Associated Molecular Patterns (DAMPs)

Sina Barghahn1, Harry Brumer2 and Volker Lipka1

1. Georg-August University Göttingen, Albrecht-von-Institute for Plant Sciences, Department of Plant Cell Biology, D-37077, Germany

2. Michael Smith Laboratories and Department of Chemistry, University of British Columbia, Vancouver, British Columbia V6T 1Z4, Canada

Plants are constantly exposed to a wide array of potential pathogens having different infection strategies and lifestyles. The plant cell wall is the first structural barrier that potential microbes need to overcome in order to infect the plant. The cell wall controls the entry success of pathogens into the host tissue and undergoes massive cell wall remodelling including callose deposition and lignification upon pathogen attack. Furthermore, the cell wall can serve as source for Danger Associated Molecular Patterns (DAMPs). DAMPs are host derived molecules that can activate immune responses. DAMPs are likely to be generated upon degradation of cell wall polysaccharides by cell wall hydrolyzing enzymes (CWHEs). Obligate biotrophic fungi, like powdery mildews, as well as hemibiotrophic fungi, like Verticillium spec. actively invade their host plants. In planta transcriptome data of the barley powdery mildew Blumeria graminis f.sp. hordei and the broad host range pathogen V. dahliae revealed that these two fungi produce distinct repertoires of CWHEs during the infection process that are likely to be essential for host plant invasion and colonization. On the other hand, it is conceivable to postulate that the activity of CWHEs on non-host or resistant Arabidopsis plants may lead to production of DAMPs that are recognized by cognate plant Pattern Recognition Receptors (PRRs) and thus trigger efficient immune responses. The major goals of this project are the identification and characterization of CWHEs of Bgh and Verticillium that are produced and required during the infection process as well as the identification of novel plant carbohydrate DAMPs and the corresponding plant perception machinery. Sina Barghahn [email protected]

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138-BS: Global insights into clubroot disease regulated genes in Arabidopsis root and shoot by RNA-Seq Solmaz Irani1, Nagabhushana Nayidu1, Brett Trost 2, Matthew Waldner2, Anthony Kusalik2, Christopher Todd1, Yangdou Wei1, Peta Bonham-Smith1* 1. Department of Biology, University of Saskatchewan, Saskatoon, SK 2. Department of Computer Science, University of Saskatchewan, Saskatoon, SK Clubroot is a plant disease that is caused by the obligate parasite Plasmodiophora brassicae. It is one of the most severe diseases of Brassicaceae as the pathogen induces the formation of galls in root tissue and disrupts the uptake of water and nutrients by the plant. Previous studies on P. brassicae infection have mainly focused on clubroot development or the pathogen life cycle in host roots. The aim of this study was to better understand the molecular basis of changes in above and below ground tissues of the host, Arabidopsis thaliana, during clubroot establishment and progression. Transcriptome analysis (RNA-seq) of roots and shoots was conducted at 17, 20 & 24 days post inoculation (dpi) with P. brassicae resting spores. Transcript levels of genes involved in the metabolism of cell wall compounds, hormones and a number of secondary metabolites changed in both infected shoot and root. Genes involved in lipid and glycolysis pathways were mainly downregulated in shoot and upregulated in root. Genes involved in the metabolism of jasmonic acid and shikimate pathway metabolites were commonly regulated in shoot and root. These whole-plant molecular responses to clubroot progression may prove to be critical for improving clubroot-associated breeding programs in the future.

Presenting author: Solmaz Irani [email protected]

139-BS: The Distribution of Laboulbeniomycetes - Obligate ectoparasitic ascomycetes in India

Surinder Kaur

Department of Botany, SGTB Khalsa College, University of Delhi, Delhi 110007, INDIA Laboulbeniomycetes, an independent class of the phylum Ascomycota consists of obligate exoparasites of true insects and other arthropods, but mites & millipedes have also been reported as hosts. These fungi usually penetrate the outer layer of the integument of the host and rarely cause disease symptoms. In fact they are too small to be noticed by mycologists and entomologists. Morphologically these fungi are different from other fungi in lacking any prominent mycelium. The thalli are small and of determinate growth bearing antheridia and perithecia on a receptacle with appendages. We conducted Extensive study of these fascinating and interesting fungi and reported about 22 species from India. As far as their distribution and occurrence in India is concerned, they are poorly studied here. We therefore carried out detailed study on ecology of these fungi along with taxonomy. The percent frequency of occurrence of these fungi on insects collected from different habitats was compared. Work was also carried out on development of these fungi and complete life cycle of these fungi was studied. The work has been carried out in the hope that this group of fungi does not remain unknown in the country and studies are being done to explore and report more species from India and on the phylogenetic relationships of these fungi,

[email protected]

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140-BS: Overexpression of AtGolS3 and CsRFS in Poplar Enhances ROS Tolerance and Systemically Represses Defense Response to Leaf Rust Disease Jonathan LA MANTIA1, 2 Faride UNDA3 Carl J. DOUGLAS4, Shawn D. MANSFIELD3, and Richard C. HAMELIN1, 5 1 Department of Forest and Conservation Sciences, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 2 United States Department of Agriculture, Wooster, Ohio 44691, USA 3 Department of Wood Science, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 4 Department of Botany, University of British Columbia, Vancouver, BC V6T 1Z4, Canada 5 Institut de Biologie Intégrative et des Systèmes, Université Laval, Québec, G1V 0A6, Canada Plants have developed a large selection of chemical, structural and protein-based defense strategies designed to respond to invading organisms and mitigate potential damage. It has been suggested that plant defense against biotrophic pathogens is mainly due to the initiation of programmed cell death in the host, and the activation of salicylic acid-dependent pathway. In contrast, during necrotrophic pathogens attack, jasmonic acid and ethylene signalling activate the defense responses. Myo-inositol and galactinol have recently been shown to participate in the defense responses for biotrophic and necrotic pathogens, respectively. To investigate the function of these compounds on defense response, we examined transgenic hybrid poplar mis-regulating galactinol synthase (GolS) (both overexpression and RNAi-mediated suppression) and overexpressing a raffinose synthase (RFS) gene challenged with the biotrophic pathogen Melampsora aecidiodes. Transgenic trees overexpressing GolS and RFS exhibiting elevated cellular concentrations of galactinol and raffinose, respectively, were more susceptible to leaf rust infection. The inoculated wild-type leaves showed reduced galactinol concentrations and repressed the expression of two endogenous GolS genes compared to un-inoculated wild-type leaves prior to the up-regulation of NON-EXPRESSOR OF PR1 and PATHOGENESIS-RELATED1. Transgenic leaves clearly showed repression of genes participating in calcium influx, phosphatidic acid (PA) biosynthesis and signalling, and salicylic acid (SA) signalling. Furthermore, transgenic trees revealed enhanced tolerance to H2O2 and up-regulation of antioxidant biosynthetic genes. These results suggest that the overexpression of GolS and RFS antagonizes the defense response to poplar leaf-rust disease through repressing reactive oxygen species and attenuating calcium and phosphatidic acid signaling events that lead to SA defense. Faride Unda [email protected] 141-BS: Clonostachys rosea ’omics profiling indicates roles for secondary metabolites in its antifungal activity against Fusarium graminearum

Zerihun A. Demissie1, Miroslava Cuperlovic-Culf2, Yifang Tan3 and Michele C. Loewen1

1 Aquatic and Crop Research and Development, National Research Council of Canada, Ottawa, ON 2 Information and Communication Technologies, National Research Council of Canada, Ottawa, ON 3 Aquatic and Crop Research and Development, National Research Council of Canada, Saskatoon, SK Clonostachys rosea strain ACM941 is a fungal bio-control agent patented in Canada against the FHB disease causative agent Fusarium graminearum. Although the molecular and biochemical basis are yet to be fully resolved, previous studies have suggested that C. rosea secretes Fusarium growth inhibitors when grown in liquid medium. To gain insight into the genetic and metabolic factors contributing to this, we developed transcriptomic (RNAseq) and metabolite profiles of C. rosea in response to deoxynivalenol (DON) and Fusarium spent media treatment. Our RNAseq analysis revealed that of the 19,064 contigs, 1,444 and 1,792 were differentially regulated by DON and Fusarium spent media, respectively. More than half of these contigs (753 and 955, respectively) were up-regulated, notably with annotations suggesting enhancement of polyketide (PK) and non-ribosomal peptide (NRP) secondary metabolites precursor synthesis along with up-regulation of their corresponding synthases. Also notable were four ABC transporters upregulated in response to Fusarium spent media. Further analysis showed that the PKSs and NRPSs belong to three gene clusters that include ABC transporters and other genes known to tailor secondary metabolites. The RNAseq transcript abundance was further validated using quantitative rtPCR. Preliminary analysis of our metabolic profiling data was also found to corroborate the RNAseq results, showing preferential enrichment

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of amino acids in liquid media where C. rosea was treated with Fusarium spent media. Taken together, our result shows that C. rosea heavily utilizes its secondary metabolism related genetic repertoire in the fight against Fusarium graminearum. Name: Zerihun Demissie; E-mail: [email protected]

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Cell Biology I Posters (142-‐CB to 151-‐CB) Wednesday July 5, FSC Atrium 11:20am -12:30pm 142-CB: SMP domain as indicator of membrane contact site tethers in Arabidopsis thaliana Francisco BENITEZ-FUENTE1*, Makoto SHIRAKAWA2, Eun-Kyoung LEE1, Abel ROSADO1 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada. Membrane contact sites (MCS) are regions where the membranes of two different organelles are in close apposition. In this microdomains, the membranes are not fused due to the presence of protein complexes working as tethers. Besides its structural function, membrane contact site tethers play roles in lipid and Ca2+ transport. These regions are widely studied in yeast and mammals, but little information is available in plants. A common feature in some membrane contact site tethers in yeast and mammals is the presence of a lipid and protein-binding domain, the synaptotagmin-like mitochondrial lipid-binding protein (SMP) domain. Recently, new MCS tethers containing the SMP domain have been identified in Arabidopsis thaliana, suggesting that this domain could also be involved in plant MCSs. We have used the sequence of the SMP domain in phylogenetic studies, finding putative tethers in Arabidopsis thaliana. Transgenic lines expressing the putative tethers transcriptionally attached to the green fluorescent protein show an endoplasmic reticulum (ER)-localized puncta pattern in the equatorial ER and a reticulate pattern in the cortical ER. This pattern has already been showed in others plasma membrane (PM)-ER tethers in plants, suggesting the putative tethers could be localized at PM-ER contact sites. Francisco Benitez-Fuente [email protected] 143-CB: The ARK2 kinesin prevents cell file rotation and root skewing in a microtubule dependent process Jorge A. HOLGUÍN CRUZ, Laryssa S. HALAT, Geoffrey WASTENEYS Department of Botany, The University of British Columbia, Vancouver, BC Certain kinesin motor proteins function in maintaining microtubule (MT) dynamics, which ensures appropriate cell growth and morphogenesis. Defects in MT array organization and dynamics have been correlated with epidermal cell file rotation and root skewing phenotypes. In Arabidopsis thaliana, the ARMADILLO-REPEAT KINESIN 2 (ARK2) is thought to regulate MT dynamics, given that a null mutant (ark2-1) displays leftward root skewing. Given that its paralogue, ARK1, has been shown to induce MT depolymerization in root hairs, we hypothesized that ARK2 also functions as a MT catastrophe factor. To test this, we applied low concentrations of the MT-disrupting drug oryzalin to the ark2-1 mutants. Similar to the rescue of the crooked root hair phenotype of ARK1 loss-of-function mutants by such treatments, 0.1 ⎧M oryzalin dramatically reduced root skewing in the ark2-1 mutant. However, using live cell imaging we found that the MT velocity and catastrophe frequency in hypocotyl cells of the ark2-1 mutant did not differ from that of wild type. MT array orientation in the ark2-1 mutant was also normal in the root elongation zone where ARK2 is expressed. Furthermore, epidermal cell file rotation was detected in the root differentiation zone of the ark2-1 mutant but not in the root elongation zone where it was expected to occur. These results indicate that the root skewing observed in ark2-1 mutants is not dependent on changes in MT orientation or cell file rotation in the root elongation zone. Jorge A. Holguín Cruz [email protected]

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144-CB: Where’s COBRA? An Affinity-Tagging Approach to Understand COBRA’s Function in Cellulose Synthesis. Karlson Pang1*, Charles Hocart2, Miki Fujita1, Geoff Wasteneys1 1. Department of Botany, University of British Columbia, Vancouver, BC 2. Research School of Biology, Australian National University, Canberra, Australia Synthesis of cellulose microfibrils is essential for maintaining the unidirectional expansion of plant organs such as roots and shoots. COBRA (COB), an essential protein that is highly expressed during rapid growth, plays an as-yet-undefined role in cellulose biosynthesis. With a cellulose-binding N-terminal domain and a GPI-anchor at its C-terminus, COB presents a challenge when designing fluorescent reporter fusion proteins to track its distribution in cells. So far only one reporter construct in which the 28 kDa yellow fluorescent protein is inserted at an interior site (COB-cYFP) is able to generate viable seedlings when expressed in the lethal cob-4 null allele, but this phenotype rescue is only partial. With COB-cYFP’s reliability as a reporter in question, we decided to tag COB instead with a smaller affinity tag. We designed a 6x-histidine-tagged COB (HisCOB) construct that, when stably expressed in Arabidopsis thaliana, fully complemented the cob-4 null mutant’s cellulose-deficient and seedling-lethal phenotype. Immunofluorescence microscopy showed HisCOB to be predominantly distributed along cortical microtubules and localized to Golgi bodies in agreement with the distribution patterns obtained in living cells with the COB-cYFP line. This finding indicates that the COB-cYFP is an accurate reporter, and that COB is indeed associated with compartments that track along cortical microtubules. This finding is in stark contrast to - and brings into question - previous immunofluorescence and immunogold-transmission electron microscopy studies in which COB was predominantly distributed to the cell wall. Karlson Pang [email protected] 145-CB: Everything in moderation: The microtubule-associated CLASP balances division and differentiation in the plant root meristem Laryssa S. HALAT1*, Geoffrey O. WASTENEYS1 1. Department of Botany, University of British Columbia, Vancouver, BC Meristems are specialized regions containing undifferentiated cells that enable plant growth to be sustained, in some cases for centuries. In Arabidopsis thaliana, the microtubule-associated protein CLASP has been identified as a controller of meristem size through its role in tethering endosomal sorting vesicles to microtubules, which fosters the recycling of the auxin transporter PIN2 and the brassinosteroid (BR) hormone receptor BRI1 to the plasma membrane. In mutants lacking CLASP expression, PIN2 and BRI1 activities are consequently reduced, and hormone signalling is perturbed. Intriguingly, CLASP’s expression is down-regulated by the BR signalling pathway, through the binding to CLASP’s promoter of two BR-activated transcription factors, BZR1 and BZR2. Based on this, we hypothesized that mutating the CLASP promoter such that it is no longer targeted by BZR1/2 would impact meristem development. Using the mutated promoter to drive expression of a green fluorescent protein-CLASP reporter in the transcript-null clasp-1 mutant, we observed fewer cells in the meristematic region compared to wild type, possibly owing to enhanced hormone receptor recycling. We confirmed that with the mutated promoter, CLASP expression is higher than normal and less affected by exogenous BR application, and that the GFP-CLASP protein is more abundant and distributed along the full length of microtubules. In addition, we found that the BR-insensitive GFP-CLASP plants were hypersensitive to osmotic stress. These findings suggest that feedback signalling between BR and CLASP is critical for plants to maintain cells in a proliferative state and to respond to abiotic stress. Laryssa Halat [email protected]

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146-CB: Is COBRA the target of the cellulose biosynthesis inhibitor Thaxtomin A?

Marcus Woodley1, Miki Fujita1, Fatima Awwad2, Nathalie Beaudoin2, Geoffrey Wasteneys1

1. Department of Botany, University of British Columbia, Vancouver, BC Canada 2. Centre Sève, Université de Sherbrooke, Sherbrooke, QC, Canada Thaxtomin A (TA), a toxin produced by the bacterial pathogen Streptomyces scabies, is a key component of common scab disease in potatoes. The specific target and mechanism by which TA enables the pathogen to infect its host is unknown, though TA has been demonstrated to inhibit cellulose production and quality. To determine how TA affects cellulose biosynthesis we used chemical genetics and live cell imaging strategies. After testing TA’s effects on a number of Arabidopsis thaliana mutants with compromised cellulose synthesis, we noted an abnormal response to TA in cobra-1 (cob-1) mutants. When grown on high sucrose media, cob-1 mutants have reduced cellulose production and roots undergo radial swelling. TA treatments, which cause cob-1-like symptoms to the wild type, partially rescued the growth defects of the cob-1 mutant. Since COBRA is a membrane-associated GPI-anchored protein and highly co-expressed with primary cell wall cellulose synthase enzymes (CesAs), we used Total Internal Reflection Fluorescence microscopy to determine how TA affects the movement and behaviour of Cellulose Synthase Complexes (CSCs). Synthesis of cellulose microfibrils by CSCs can be measured by tracking fluorescently tagged CesA subunits, many of which are seen in close association with cortical microtubules. TA treatment caused a significant reduction in the CSC speed, and generated aberrant movement of CSCs in regions devoid of microtubules. These results identify COBRA as a possible target of TA activity, and suggest that this interaction compromises the normal catalytic activity of the CSCs, thereby making plant cell walls vulnerable to pathogen attack.

Marcus Woodley [email protected]

147-CB: MOR1 on the move: Using FRAP analysis to describe MOR1 protein dynamics on moving microtubule ends

Ron BLUTRICH1, Laryssa S. HALAT1, Geoffrey O. WASTENEYS1

1. Department of Botany, University of British Columbia, Vancouver, BC

Microtubules (MTs) are polymers that are critical for plant development, the dynamics and organization of which are modulated by other proteins. In the plant Arabidopsis thaliana, MICROTUBULE ORGANIZATION 1 (MOR1) is a plus-end tracking protein that plays a role as a MT polymerase. While the indispensible role of MOR1 in promoting MT growth is known, its mechanism of function and MT association remains unclear. To determine the dynamicity of MOR1 on MTs, we used fluorescence recovery after photobleaching (FRAP) in conjunction with mathematical modeling. To visualize live cells expressing MOR1, a MOR1pro:MOR1-3xYpet construct was transformed into wild-type plants as well as a T-DNA insertion line, mor1-23. FRAP was performed by photobleaching multiple MOR1 punctae within a defined region of interest (ROI). A model that adjusts for photobleaching most accurately represented recovery parameters for MOR1. Since the ROI encompassed multiple MOR1 punctae, we decided to test if there was a correlation between starting distance from the centre of the ROI and recovery dynamics. The position of MOR1 within the ROI had a significant effect on the recovery rate and mobile fraction, which prompted us to obtain recovery measurements for individual MOR1 punctae. When comparing dynamics between the wild-type and mor1-23 genotypes, MOR1 had a decreased affinity for the MT in the wild-type background, suggesting that the endogenous, unlabeled MOR1 protein was able to outcompete the fluorescently tagged protein for binding sites. Our results describe in-depth imaging and modeling techniques that can be used to assess protein dynamics on growing MTs.

Ron Blutrich [email protected]

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148-CB: Mechanisms influencing the polar distribution of pectin and cell wall proteins in seed coat epidermal cells of Arabidopsis Yi-Chen LEE1*, Erin GILCHRIST2, Allen Yi-Lun TSAI3, George HAUGHN1 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada 2. Anandia Laboratories, Vancouver, BC, Canada 3. Graduate School of Science and Technology, Kumamoto University, Chuo-ku, Kumamoto, Japan

During differentiation, Arabidopsis seed coat epidemal cells deposit large amounts of pectin-rich mucilage in a

polar manner to the outer tangential side of the cell forming a large apoplastic pocket. MUM2 is a β-galactosidase that modifys pectin in the mucilage. Using an engineered version of MUM2 fused to yellow flourescent protein (YFP), the distribution pattern of MUM2 in the epidermal cell was determined. MUM2-YFP is found to preferentially accummulate in the mucilage pocket concomitantly with pectin deposition. The amino acid sequence of MUM2 is not required for the secretion to the pocket. Rather, the polar distribution of MUM2-YFP is caused by a rearragement of the secretory pathway that appears to target all secretion to the outer tangential side of the cell. At the end of mucilage synthesis, the fluorescence of MUM2-YFP rapidly disapears from the mucilage pocket. The results of western blot analyses showed that the amount of MUM2-YFP decreases, suggesting that the disapearance of MUM2-YFP signal is due to protein degradation. A loss-of-function mutation in the gene encoding ASPG1, a protease that has been detected in mucilage from mature seeds, resulted in a delay of MUM2-YFP degradation, suggesting that ASPG1 is responsible, at least in part, for the degradation.

Yi-Chen Lee [email protected] 149-CB: Cytoskeleton independent endoplasmic reticulum-plasma membrane contact site dynamics upon ionic stress in Arabidopsis EunKyoung Lee1, Steffen Vanneste2,3, Francisco Benitez-Fuente1, Jiri Friml2,3, Abel Rosado1 1. Department of Botany, Faculty of Sciences, University of British Columbia, Vancouver, Canada, V6T 1Z4 2. Department of Plant Systems Biology, VIB, Ghent, Belgium 3. Department of Plant Biotechnology and Bioinformatics, Ghent University, Ghent, Belgium

ER-RM contacts occur in all eukaryotic cells and control intracellular Ca2+ dynamics and lipid homeostasis. In plant, ER-PM contact sites (EPCSs) are essential for the regulation in response to developmental signals and environmental stresses. However, the distributions and dynamics of EPCSs under stresses remain unclear. Plant EPCSs associate with the cytoskeleton, keeping them stable through an EPCS protein complex including cytoskeleton-binding proteins. Arabidopsis Synaptotagmin 1 (SYT1) is a well-known tether between the ER and PM. We found that the localization and dynamics of SYT1-GFP were largely reorganized upon ionic stress. The ionic stress caused by salt treatment induces for cortical EPCS tubules to be linked more each other, resulting in the reticulated structures on the cortical ER. This stress induced-SYT1 reorganization is not dependent upon cytoskeletons. In addition, pharmacologically increased PI(4,5)P2 levels also induce the reorganization of SYT1 protein being reminiscent of those by ionic stress. This result suggests that the phospholipid binding-C2 domain of SYT1 can be mainly involved in the rearrangement of SYT1 localization upon ionic stresses. Together, during ionic stress exposure, the reorganization of cortical EPCS is likely a common response in leaf epidermal cells.

EunKyoung Lee [email protected]

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150-CB: COBRA, a protein essential for plant directional cell growth, is predominantly associated with microtubules and Golgi Bodies in elongating cells of Arabidopsis thaliana Miki Fujita1, Karlson Pang1, Charles, Hocart2, Geoffrey O. Wasteneys1 1Department of Botany, University of British Columbia, Vancouver, Canada 2Research school of Biology, Australian National University, Canberra, Australia COBRA (COB) is a glycosylphosphatidylinositol (GPI)-anchored protein known to be involved in cellulose biosynthesis in rapidly elongating plant cells. Loss-of-function cobra mutants have swollen roots and reduced cellulose levels. COB has a N-terminal cellulose-binding domain but how COB functions in cellulose biosynthesis is still unknown. In this study, we generated transgenic lines that express COB-citrineYFP (cYFP), which rescue the seedling-lethal phenotype of the cob-4 null mutant. Although the roots of COB-cYFP/cob-4 were short at the seedling stage, cellulose content was restored to the wild-type level. In contrast to previous immunolabelling studies showing COB’s abundance in cell wall, COB-cYFP was found in small puncta associated with the microtubules. COB-cYFP was also abundant in Golgi bodies and we observed periodic removal of COB-cYFP puncta from microtubules by Golgi complexes. Microtubule-associated COB-YFP puncta moved at two velocities: relatively fast movement consistent with kinesin motility and much slower velocities similar to those recorded for cellulose-synthase-complexes (CSCs). Using near total internal reflection fluorescence (TIRF) microscopy, we found a strong correlation with the slow movement along microtubules of COB-YFP puncta and the cellulose-synthase interacting protein 1 (CSI1/POM2), which is involved in linking CSCs to microtubules. Unlike the COB-cYFP puncta, CSI1/POM2 was not removed from microtubules by trans-Golgi complexes. Our results indicate that microtubules are a major hub for COB trafficking and that the turn over of microtubule-associated COB-cYFP puncta is distinct from that of CSCs. Miki Fujita: [email protected] 151-CB: Regulation of Bilateral Symmetry in Stomata by the Leucine-Rich Repeat Receptor-Like Kinase Mustaches Sandra KEERTHISINGHE1, Fred SACK1 and Geoffrey WASTENEYS1 1. The University of British Columbia Stomata are specialized plant epidermal structures consisting of two bilaterally symmetrical guard cells surrounding a pore. Co-ordinated guard cell movement controls pore opening, thereby regulating the gas exchange required for photosynthesis. Stomata develop via a pathway defined by two key divisions, an asymmetric division, and a symmetric division, which initiates stomatal bilateral symmetry. This initial symmetry is then maintained by the subsequently occurring pore and guard cell morphogenesis. The Leucine-Rich Repeat Receptor-Like Kinase MUSTACHES (MUS) is required to enforce bilateral symmetry post-symmetric division. Mutants in mus display pore and guard cell morphogenesis defects, as well as defects in microtubule array organization and polarity of microtubule movement. Further characterization of MUS function, utilizing time-lapse studies, demonstrated that MUS maintains bilateral symmetry by enforcing the symmetrical positioning of stomatal Microtubule Organizing centres (MTOCs) at the centre of the stoma. Time lapse studies also indicated that delocalization of MTOCs away from the stomatal center occurred before, and is likely responsible for, the subsequently occurring alterations in microtubule polarity and defective guard cell morphogenesis observed in mus. Finally, double mutant analysis indicated that the formation and placement of the stomatal MTOC was dependent on MUS, and the microtubule associated proteins CLIP-170 ASSOCIATED PROTEIN (CLASP) and NIMA-RELATED KINASE6 (NEK6). These findings corroborate the existence of a stomatal-specific cortical MTOC, and the importance of the polarity of microtubule growth in contributing to stomatal morphogenesis. Additionally, to our knowledge, this work is the first to describe potential mechanisms and genetic relationships that influence MTOC formation in stomata. [email protected]

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Nutrients and Metabolism Posters (152-‐NM to 154-‐NM) Wednesday July 5, FSC Atrium 11:20am -12:30pm 152-NM: Chlamydomonas reinhardtii as a platform to study nitrogen sensation and transcriptional responses in the green lineage Jacob A. MUNZ1*, Sunjoo JOO1, Jae-Hyeok LEE1 1. Department of Botany, University of British Columbia, Vancouver, BC Nitrogen (N) is a fundamental macronutrient necessary for nucleotides and amino acids, yet most organisms rely on external N-sources in their immediate environment. Nitrogen utilization efficiency is of fundamental importance to crop yields, however, the precise mechanisms by which photosynthetic eukaryotes sense their overall N-status and trigger appropriate responses remain unknown. The unicellular green algae Chlamydomonas reinhardtii (Chlamydomonas) provides an effective system for finely targeted studies within major processes like N-sensation without the complications of body plan or sophisticated culturing systems. Additionally, Chlamydomonas has clear N-status responses: gametogenesis, photosynthetic electron transport chain degradation, starch/lipid accumulation, and gene regulation. Transcriptomics studies have found rapid changes in gene expression after removal of the N-source suggesting sensor-mediated signaling mechanisms in the control of N-starvation responses that largely remain elusive. We have generated an insertional mutant library harboring a reporter construct driven by a promoter responding to N-starvation. We selected for mutants with the N-starvation responsive reporter either constitutively active or no longer responsive to N-starvation. Screening of >10,000 mutants has revealed 43 mutants with defects controlling one or more of the aforementioned N-status responses. Preliminary characterization of a mutant (ns102) unable to activate the N-starvation responsive promoter found that it lacks the ability to activate N-starvation responsive genes or to initiate the gametogenesis program indicating defective sensing or responses to N-starvation. Our collection of mutants will provide insight into how photosynthetic eukaryotes sense and respond to varying N-levels opening avenues to engineer organisms with desirable properties. Jacob Munz [email protected]

153-NM: Combined transcroptomic and metabolomic approaches provide new insights into C/N partitioning in roots of Arabidopsis thaliana.

Shrikaar Kambhampati*1,2, S. Pandurangan1,2, J.B. Renaud2, R.S. Austin2, M.W. Sumarah2, F. Marsolais1,2.

1. Department of Biology, Western University, London, Ontario

2. London Research and Development Center, Agriculture and Agri-Food Canada, London, Ontario

Balance between carbon and nitrogen metabolism is a requirement for the sustained growth of organisms. In plant leaves, this balance is achieved by inter-relationships between photosynthesis, respiration and amino acid metabolism in a photoperiod dependent manner. The GS/GOGAT cycle is a well understood mechanism in plants known to serve as a cross-road between carbon and nitrogen metabolism. Non-photosynthetic tissues (e.g., roots, germinating seeds), however, lack a sufficient supply of carbon skeletons under high nitrogen conditions and hence may resort to other mechanisms, along with the GS/GOGAT cycle, to achieve the desired carbon/nitrogen balance. Considering the importance of asparagine as a major storage form of nitrogen, this study elucidates carbon and nitrogen partitioning within Arabidopsis roots upon asparagine treatment. Here, we propose a potential role for the GAT1_2.1 enzyme in hydrolyzing glutamine to glutamate which can serve as a carbon skeleton for channeling carbon to the TCA cycle, under high nitrogen conditions. Transcriptome analysis revealed a 4.3 fold upregulation of a class I glutamine amidotransferase, GAT1_2.1; GAT1_2.1 was shown to be highly responsive to nitrogen levels and has a root specific expression in Arabidopsis. Metabolite profiling data further strengthened the transcriptome data and suggest a major reprogramming of C and N metabolites to sustain TCA cycle.

Shrikaar Kambhampati [email protected]

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154-NM: Effects of Copper Limitation on the Photosythetic Proteome of the Open Ocean Diatom Thalassiosira oceanica Anna A. HIPPMANN1, Nina SCHUBACK1¶, Kyung-Mee MOON2¶, John P. McCROW3¶, Andrew E. ALLEN3, Leonard J. FOSTER2, Maria T. MALDONADO1, Beverley R. GREEN4 1.Department of Earth, Ocean, and Atmospheric Sciences, University of British Columbia, Vancouver, BC, Canada 2.Department of Biochemistry & Molecular Biology, University of British Columbia, Vancouver, BC, Canada 3.Department of Microbial & Environmental Genomics, J. Craig Venter Institute, La Jolla, CA, USA 4.Department of Botany, University of British Columbia, Vancouver, BC, Canada Diatoms account for a large fraction of carbon fixation in the ocean, but how they cope with low Cu levels in the open ocean is largely unknown. We show that in response to low Cu there is a comprehensive restructuring of the photosynthetic apparatus in the diatom Thalassiosira oceanica (CCMP1003), at both the physiological and proteomic level. The result is a shift from light harvesting for photochemistry — and ultimately for carbon fixation — to photoprotection, reducing carbon fixation and oxygen evolution. Concomitant with decreases in photosynthetic efficiency (Fv/Fm), carbon fixation, and oxygen evolution, there are increases in the antennae absorption cross section (σPSII) and non-photochemical quenching (NPQ). At the proteomic level, low Cu induces a significant four-fold reduction in the Cu-containing photosynthetic electron carrier plastocyanin, which causes a bottleneck in the photosynthetic electron transport chain (ETC), ultimately leading to substantial stoichiometric changes. Namely, a two-fold reduction in both cytochrome b6f complex (cytb6f) and photosystem II (PSII), no change in the Fe-rich PSI and a 40- and 2-fold increase in proteins potentially involved in detoxification of reactive oxygen species (ferredoxin and ferredoxin:NADP+ reductase, respectively). We identified 48 light harvesting complex (LHC) proteins in the publicly available genome of T. oceanica and provide proteomic evidence for 33 of these. The change in the LHC composition within the antennae in response to low Cu underlines the shift from photochemistry to photoprotection in T. oceanica. Beverley Green [email protected]

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Sexual Reproduction Posters (155-‐SR to 160-‐SR) Wednesday July 5, FSC Atrium 11:20am -12:30pm 155-SR: The temporal regulation of Flavanoid and Reactive oxygen species in the stigma regulate self-pollen rejection responses in Kale Abhinandan Kumar1,3*, Xingguo Lan1,2,3, Yuhua Li2, Marcus A. Samuel1 1. Department of Biological Sciences, University of Calgary, Calgary, AB, T2N 1N4, 2. Canada College of Life Sciences, Northeast Forestry University, Harbin, 150040 Heilongjiang, P R China, 3. These authors contributed equally to this article. Self-incompatibility (SI) in hermaphroditic angiosperms is a complex mechanism of mate selection/rejection during sexual reproduction. The landing of a self-pollen on the stigma of the Brassicaceae family (kale, canola, cabbage) triggers a haplotype-specific rejection of the pollen caused by a ligand-receptor (SCR/SRK) mediated intracellular signaling cascade. Except for a few identified downstream targets, knowledge of proteins involved during this response and their temporal regulation has remained elusive. Here we report 107 differentially expressed proteins (DEPs) in the developing stigma (Stage 1- Stage 5) of kale using 2D-DIGE and MALDI-TOF-TOF MS/MS. From the analysis of these DEPs, we have identified a progressive downregulation of the flavonoid-related proteins with concomitant upregulation in the levels of reactive oxygen species (ROS). In-vivo confocal imaging of the ROS levels in the stigmas using DCFH2-DA revealed an increase in ROS during stigma maturity. Our ROS quenching studies indicate that ROS accumulation is not required during SI response while it was necessary for successful pollination to occur. Supplementing mature stigmas with kaempferol before flower opening mimicked the ROS quenching studies suggesting that flavonoids could influence ROS levels in the stigmas. Interestingly, kaempferol treatment led to the breakdown of SI response in mature stigmas suggesting an accumulation of flavonoids could prevent the full manifestation of SI response in mature flowers. These findings suggest that ROS and flavonoid pathways play distinct and overlapping functions during compatibility and SI responses in kale and have likely coevolved as an outcome of selective pressure to maintain vigor while avoiding unfavorable interactions at the stigmatic surface. Abhinandan Kumar- [email protected] 156-SR: Investigating the role of autophagy in Arabidopsis self-incompatibility Hayley NELLES1*, Daphne R. GORING1,2 1. Department of Cell and Systems Biology, University of Toronto, Toronto M5S 3B2, Canada 2. Centre for the Analysis of Genome Evolution and Function, University of Toronto, Toronto M5S 3B2, Canada In the flowering plants, fertilization is controlled by a series of interactions between pollen and pistil. Members of the Brassicaceae have dry stigmas, allowing a plant to strictly regulate pollen acceptance through the selective hydration of compatible pollen grains. Most species within this family have acquired an outcrossing mechanism, known as self-incompatibility (SI), where self-pollen is rapidly rejected at the stigma surface. SI is achieved by disrupting the vesicular trafficking of stigmatic compatibility factors to the pollen contact site, thus preventing the germination of incompatible pollen. While the upstream SI signaling components, SCR and SRK, have been characterized in Arabidopsis lyrata, the downstream signaling events need further investigation. In this study, we have transformed the inbreeding species A. thaliana with three genes from A. lyrata to establish a stable SI line. Stigmas from the SI line showed a specific rejection of self-pollen, resulting in a substantial reduction to seed set. We plan to further investigate the genetic requirements of SI and the putative role of autophagy in this mechanism. A GFP:ATG8 marker was introduced into SI lines and preliminary confocal work has revealed autophagosome formation in self-pollinated stigmatic papillae. The requirement for autophagy in SI will be further explored using autophagy mutant lines (atg 5-1, atg7-2). Hayley Nelles [email protected]

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157-SR: Interaction of ribosome biogenesis factors HRR25-LTV1 Kumarakurubaran SELVARAJ *, Peta BONHAM-SMITH Department of Biology, University of Saskatchewan, Saskatoon, SK Ribosomes are ubiquitous cellular nano-machines that translate the genome into the proteome of all living cells. Ribosome biogenesis is often linked to cellular homeostasis and growth rate. Ribosome biogenesis is well described in bacteria, yeast and mammals, however, little is known in plants. In yeast, ribosome biogenesis requires multiple rRNA processing steps in concurrence with ~250 ribosome biogenesis factors (RBFs). In this process, HRR25 is an essential gene encoding a casein kinase1-like (CK1L) protein that interacts with the RNA helicase LTV1. This HRR25-LTV1 complex regulates the incorporation of small subunit ribosomal proteins RPS3 and RPS10 that line the mRNA entry channel of the 40S small subunit. HRR25-TIF6 interactions regulate the 60S large subunit association for translation of incoming mRNA. Both these interactions are key ribosome maturation events. In Arabidopsis thaliana, 14 CK1L isoforms (1-13, 9α- β) have been reported as plasmodesmata-associated kinases. Phylogenetic analysis of these CK1L isoforms shows five of the 14 (including a pseudogene) closely cluster with yeast HRR25p. Yeast two hybrid studies show differential association of each of the four AtHRR25A-D isoforms with the single copy of AtLTV1. Differential, tissue-specific expression of the four isoforms suggests that there may actually be tissue-specific, developmental-specific and/or stress-specific AtHRR25A-D: LTV1 interactions in Arabidopsis. [email protected]

158-SR: Bumps in the Road: Presence and distribution of papillae may demonstrate the dual nature of pollination in Arceuthobium americanum Matthew MCISAAC1* 1. Department of Biological Sciences, Thompson Rivers University, BC Dwarf mistletoes (Arceuthobium spp.) are dioecious angiosperms that obligately parasitize members of the Pinaceae and Cupressaceae. The Arceuthobium genus negatively impacts up to 3.8 million m3 of lumber crop annually, and the damage translates into billions of dollars in losses for the forestry industry. Arceuthobium americanum (lodgepole pine dwarf mistletoe) is a diminutive shrub that infects the lodgepole pine tree (Pinus contorta ssp. latifolia). Trees become infected when the pistillate plant explosively discharges its seed which, upon adherence to host tissue, develops a root-like penetration wedge that burrows through the bark towards the vascular cambium. Post-penetration, an endophytic system develops as highly-modified stem tissue (haustoria) extends into the ray parenchyma and accesses the hosts nutrient and water infrastructure. Two to five years later, the plant produces aerial shoots to begin the reproductive cycle. The Arceuthobium genus’ pollination biology has been the subject of discussion, as it presents floral characteristics indicative of both insect-pollinated (entomophilous) and wind-pollinated (anemophilous) strategies. In addition to these features, the presence of papillae (small nipple-like protuberances) may offer an additional advantage to insects attempting to access Arceuthobium’s pollination rewards (nectaries and stigmatic exudate), as these surface features may provide greater traction for visiting insects. The objective of this work is to quantitatively analyse the spatial distribution of papillae on staminate Arceuthobium americanum’s perianth using environmental scanning electron microscopy. Matthew McIsaac [email protected]

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159-SR: Pollen acceptance or rejection in the mustard family Emily Indriolo1, M. Isabella Chavez1, Jonathan Teske1, KassaDee Ketelaar2, Ian Wallace2, Darya Safavian3 and Daphne Goring3 1Department of Biology, New Mexico State University, Las Cruces, NM 2Department of Biochemistry and Molecular Biology, University of Nevada Reno, Reno, NV 3Department of Cell & Systems Biology, University of Toronto, Toronto, ON

Pollen-pistil interactions in flowering plants are tightly regulated to either accept compatible or reject self-pollen. In the Brassicaceae, the rejection of self-pollen is regulated by a signaling pathway activated by the stigma-specific S Receptor Kinase (SRK), following binding of a pollen-specific ligand, SCR/SP11. In Brassica species, downstream signaling components of the pathway have been identified such as the M Locus Protein Kinase and the ARC1 E3 ubiquitin ligase which targets the Exo70A1 subunit of the exocyst complex. Recently, it has been demonstrated that vesicle trafficking and the contents of secretory vesicles play an important role in pollen acceptance in the Brassicaceae. Progress has been made in determining this signaling pathway which includes Exo70A1. Previously, Exo70A1 was demonstrated to function in planta in vesicle delivery at sites of polarized secretion. Recently, it was shown that the exocyst complex is required for the delivery of factors to accept compatible pollen. However, many questions remain in the field as the identification of the receptor(s) that recognize compatible pollen grains through the basal pollen response that activates the transportation of secretory vesicles to the plasma membrane. To identify the receptor(s) of a compatible pollination and the signaling pathway(s) of the secretory vesicle release, we have been performing quantitative proteomics of B. rapa stigmas in compatible and self-incompatible pollinations. We are currently analyzing the data from our preliminary proteomics results to identify likely candidates in the basal pollen response. These candidates are currently being characterized in T-DNA knock-out lines to examine their potential role in compatible pollen acceptance. Our experiments will result in building a model for vesicle trafficking in the Brassicaceae. Poster presentation: [email protected] 160-SR: A functionally redundant MAPK cascade mediates stigma receptivity in Arabidopsis Muhammad Jamshed1, Subramanian Sankaranarayanan2, Abhinandan Kumar1 and Marcus A. Samuel1

1. Department of Biological Sciences, University of Calgary, Calgary AB T2N 1N4, Canada 2. Institute of Transformative Bio-Molecules (WPI-ITbM), Nagoya University, Furo-cho, Chikusa-ku, Nagoya, Aichi 464-8602, Japan Pollination is one of the key events during sexual reproduction in plants, which relies on complex molecular communication and signaling between pollen grains and stigmatic papillary cells. Successful pollination is achieved when pollen germinates on stigmatic surface and efficiently penetrates through papillary cell to reach the ovule, where it fertilizes the egg. The interplay of several pollen and stigma proteins decides the fate of successful pollination. Despite the identification of a number of pollen and stigma proteins required for successful pollination, the molecular signaling mechanism behind how these proteins coordinate pollination responses has remained elusive. Here we have identified a highly functionally redundant MAPK matrix in controlling pollination responses in Arabidopsis. Our genetic analyses demonstrate that in stigmas, five MAPK kinases (MKKs), MKK1/2/3/7/9 are required to transmit upstream signals to two MPKs, MPK3/4, to mediate compatible pollination. Compromised functions of these five MKKs in the quintuple mutant (mkk1/2/3RNAi/mkk7/9) phenocopied pollination defects observed in the mpk4RNAi/mpk3 double mutant. The extreme functional redundancy observed in the MAPK network during pollination highlights the importance of this pathway for successful pollination and reveals the complexity of the MAPK circuit. [email protected]

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Posters – Session 2 Thursday July 6, FSC Atrium 11:20am -12:30pm

Applied Biology / Global Food Security Posters (199-‐AP to 210-‐AP) 199-AP: Re-discovering Sargassum: Morphological and Molecular Identification of Sargassum (Fucales, Phaeophyceae) Species in Eastern Samar, Philippines for Natural Product Screening.

Leni G. YAP-DEJETO1, 2, Mark Angelo PONTICA1, and Marilou SISON-MANGUS2

1. Division of Natural Sciences & Mathematics, University of the Philippines Visayas-Tacloban College, Tacloban City, Philippines 2. Earth & Marine Sciences, Ocean Sciences, University of California Santa Cruz, CA.

The genus Sargassum is the most species-rich genus of the order Fucales. It is widely distributed and has been known to colonize many parts of the globe. Ecological and economic significance of this genus include being nurseries to marine life, and well-known fucoidan nutritional supplements. Sargassum is abundant and highly speciose in tropical Philippines but unutilized. Screening for natural products could boost its value. But a thorough identification of species members, in this archipelago is first needed. Morphological assessment was conducted to describe the Sargassum species in the province of Eastern Samar, Philippines, where several Sargassum species co-occur. Samples collected from nine locations were identified by gross morphological characteristics of the species such as the type of holdfast, shape and nature of its branches, vesicles, leaves, nature and distribution of cryptostomata, and nature and form of receptacle. Thirteen species were identified and their morphological characteristics were assessed. Out of the 13 species morphologically identified, the following are of first record in Eastern Samar, Philippines to date: S. baccularia, S. binderi, S. cinereum, S. hemiphyllum, S. henslowianum, S. incisifolium, S. kuetzingii, S. longifructum, S. parvivesiculosum, and S. tenerrimum. We conducted molecular identification of these distinct Sargassum species with the use of molecular markers followed by phylogenetic analysis to determine if the traditional classification scheme is consistent with their molecular identities. We genetically compared and validated the identity of these species and discuss our findings in the light of Sargassum speciation and diversification in the tropics.

Leni G. Yap-Dejeto [email protected] / [email protected]

200-AP: Foliar boron increased fruit set and pollen grain germination for peach in a subtropical climate Rayane Barcelos Bisi1, Filipe Bittencourt Machado de Souza1, Rafael Pio1, Lee Kalcsits2, Guilherme Locatelli1 , Carolina Ruiz Zambon1 1- Department of Plant Science, Federal University of Lavras, 37200-000 Lavras, Minas Gerais State, Brazil. 2- Department of Horticulture, Washington State University, Tree Fruit Research and Extension Center,

Wenatchee, WA, US.

Flower pollination and fertilization success is a major factor contributing to the overall yield potential of Prunus persica (peach). Previous research has demonstrated that some cultivars lack consistent cropping (yield variation) in subtropical regions. Boron has been reported to influence overall fruit set in horticultural crops. Here, the application of boric acid at flowering was tested as a strategy to improve yield stability for peach grown in subtropical climates. Pollen germination and fruit set was measured for two peach cultivars when treated with three concentrations of boric acid (400 mg L-1, 800 mg L-1 or 1,200 mg L-1) or a control treatment of water at budbreak. Treatments were applied to dormant shoots when flower buds were swelling on early maturing cultivars ‘Aurora 2’ and ‘Doçura 2’. Pollen grain germination and fruit set decreased in ‘Aurora 2’ but increased for ‘Docura 2’ at a concentration of 400 mg L-1 of boric acid. To evaluate the cultivar-specific response to foliar boric acid treatments, a second experiment using only the water control and 400 mg L-1 boric acid treatment was conducted using 18 cultivars. The application of 400 mg L-1 of boric acid increased fruit set on most of the cultivars tested. Whereas, pollen grain germination increased in only a few of the cultivars. The response of boron applications to peach in a subtropical climate was cultivar dependent and was not uniformly successful.

Rayane Barcelos Bisi [email protected]

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201-AP: The Effects of Seasonal Changes on Photosynthesis Rates, Transpiration Rates and Protein Levels in the Leaves of Red Maple (Acer rubrum), Red Oak (Quercus rubra) and Western Red Cedar (Thuja plicata) Amneet DHILLON1*, Jarnail CHANDI1, Santokh SINGH1 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada V6T 1Z4 The ability of plants to capture carbon dioxide looks to be a promising solution to the increasing levels of greenhouse gases in the atmosphere. However, there is conflicting evidence in regards to the relative effectiveness of different species of plants. This study aimed to compare chlorophyll, protein, photosynthesis and transpiration levels in A. rubrum, Q. rubra and T. plicata and to provide insight into which species may be the most efficient at sequestering carbon dioxide. The chlorophyll levels were measured only in A. rubrum and Q. rubra, using the CL-01 Chlorophyll Content System. Chlorophyll levels were consistently higher in A. rubrum than in Q. rubra but both species showed a dramatic decrease in early October. A. rubrum and Q. rubra showed relatively low, but consistent photosynthesis and transpiration rates throughout the study while the T. plicata had relatively higher rates with an intense peak in late July and early August. We also analyzed the RuBisCO and the Light Harvesting Complex IIb (LHC IIb) proteins in the leaves of these plants by using the Western blot technique. Both A. rubrum and T. plicata showed very high amounts of both RubisCO and LHC IIb, while Q. rubra showed very low amounts. In summary, T. plicata appears to show a higher net photosynthesis rates especially from July to October. The environmental significance and physiological mechanisms of these research findings will be discussed. Amneet Dhillon [email protected]

202-AP: Precision irrigation in nursery using wireless tensiometers Isabelle CLERMONT1, Aurélie MUNGER1, Louis-Félix NADEAU1, Blandine BULOT1, Philippe CÔTÉ1, Benjamin DOUCET1, Antoine PAQUET1, Charles GOULET1 1. Centre de recherche et d’innovation sur les végétaux, Département de phytologie, Université Laval, Québec, QC Water management is one of the most important issues in nursery plant production. Providing enough water to the plants is essential to maintain an optimal growth and has a direct impact on both plant appearance and the duration of the production cycle. Nonetheless, deciding when to irrigate is often challenging and overwatering can be detrimental both for the environment and for the production costs. To optimize water use and help in the decision making, we evaluate a new generation wireless tensiometer that measures the water available to the plant and allows precise irrigation control. Using a web interface, we were able to monitor changes in water availability to the plants in real time. This allowed precise control and an accurate assessment of the best irrigation threshold for ten reference species. There were major differences in the total volume of water used by the species regardless of their optimal threshold. For example, the Hosta used around seven times less water than the Astilbe despite growing the best at the same irrigation threshold. This difference, which can be explained in part by their contrasted growth rate, illustrated the need of creating clusters based on their overall water needs during a season. Based on these results, we tried to find the best clustering practices for a wide range of species by combining them to the reference species monitored with tensiometers. The results demonstrate how it is possible to reduce water use by making efficient clustering of species in a nursery.

Isabelle Clermont, [email protected]

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203-AP: Does haskap (Lonicera caerulea L.) benefit from nitrogen fertilization?

Julie LAJEUNESSE*, and Jean LAFOND Quebec Research and Development Center, Agriculture and Agri-Food Canada, Normandin Research Farm, Qc, Canada Haskap (Lonicera caerulea L.) seems to offer a high potential for production and in Quebec, there is a growing interest in cultivating this berry crop. Little information is available on crop management and research is needed to establish provincial recommendations. In fall 2015, a trial was established in a 4 year-old orchard to evaluate 4 cultivars (Indigo Yum, Indigo Gem, Indigo Treat, and Tundra), three nitrogen source (ammonium nitrate, urea, and calcium nitrate), and four nitrogen rates (0, 27.5, 55, 110 kg N ha-1). The experimental design was a split-split-plot where whole plot was the cultivar, subplot was the nitrogen source, and the sub-subplot was the nitrogen rate. The choice of cultivar affected significantly fruit yields. In 2016 fruits yields varied from 77.5 and 512.9 g per plant. The cultivar Indigo Gem had the highest fruit yield compare to other cultivars. Nitrogen source and rate had no significant effect on fruit production in 2016. This trial will continue in 2017. Julie Lajeunesse [email protected] 204-AP: Genome-wide analysis of cis-regulatory element structure and discovery of motif-driven gene co-expression networks in grapevine (Vitis vinifera L.)

Rodrigo Lopez Gutierrez1, Darren C. Wong1, Gregory A. Gambetta2, Simone D. Castellarin1

1. Wine Research Centre, University of British Columbia, Vancouver, BC, Canada 2. Ecophysiologie et Ge´ nomique Fonctionnelle de la Vigne, Bordeaux Sciences Agro, INRA, Universite´ de

Bordeaux, 33140 Villenave d’Ornon, France

Wine quality is affected by the accumulation of metabolites in the grape berry, a process which is modulated by a plethora of genes. Biotic and abiotic stresses strongly affect the expression of these genes. Studying the structure and regulatory mechanisms of genes provides a strong foundation for elucidating grape metabolism and predicting its response to the various stimuli. Transcription factors (TFs) act to modulate gene expression through complex cis-regulatory element (CRE) interactions. Genome-wide analysis of known plant CREs was performed for all currently predicted protein-coding gene promoters in grapevine. Many CREs such as abscisic acid (ABA)-responsive, drought-responsive, auxin-responsive, and evening elements, exhibit bona fide CRE properties such as strong position bias towards the transcription start site (TSS) and over-representation when compared with random promoters. Genes containing these CREs are enriched in a large repertoire of plant biological pathways. Large-scale transcriptome analyses also show that these CREs are highly implicated in grapevine development and stress response. Numerous CRE-driven modules in condition-specific gene co-expression networks (GCNs) were identified and many of these modules were highly enriched for plant biological functions. Several modules corroborate known roles of CREs in drought response, pathogen defense, cell wall metabolism, and fruit ripening. Comparisons with Arabidopsis suggest a general conservation in promoter architecture, gene expression dynamics, and GCN structure across species. Systems analyses of CREs provide insights into the grapevine cis-regulatory code and contribute to the knowledge on fruit metabolism and how it is modulated by biotic and abiotic factors in vineyards.

[email protected]

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205-AP: An examination of Poplar and Willow for Phytoremediation Potential of Salinization at Industrial Sites Michael BILEK1* Raju SOOLANAYAKANAHALLY1,2, Robert GUY3, Shawn MANSFIELD1 1. Department of Wood Science, University of British Columbia, Vancouver, B.C. 2. Agriculture and Agri-Food Canada, Agroforestry Development Centre, Indian Head, S.K. 3. Department of Forest and Conservation Sciences, University of British Columbia, Vancouver, B.C. Land degradation due to soil salinity is a worldwide issue, with recent conservative estimates of soil salinization affecting 10% of global landmass. Natural and anthropogenic causes of soil salinization including bedrock weathering, poor irrigation practices, and mining runoff significantly threaten productive land. Moreover, contemporary methods of remediating contaminated soils are costly and extremely involved processes, and a suitably inexpensive alternative is becoming increasingly necessary. Employing naturally salt tolerant native species to inhabit and ameliorate contaminated land is a favorable, economically viable option. Poplar and willow tree species are excellent candidates, as they are fast growing, and easy to propagate by cuttings. The aim of this project is to identify native poplar, willow, and poplar-hybrid genotypes for salt tolerance. Two sequential greenhouse trials were conducted that examined the growth response of native genotypes to varying levels of soil salinization (NaCl) in a randomized block design. Thirty-six initial genotypes were grown for 3 months in 0, 30, and 80 mM NaCl soil solutions, from which eight selected genotypes were further examined for four months at 0, 20, 40, and 60 mM NaCl. Poplar genotypes exhibited 100% mortality at treatment levels greater than 60 mM NaCl, while willow genotypes experienced an average of 14.6% reduction in height and 24.4% reduction in stem diameter at 80 mM NaCl. By planting candidate genotypes on contaminated sites, they may act as pioneer species in otherwise inhospitable environments, and consequently permit the regeneration of additional plant species and ultimately contribute to the restoration of affected sites. Michael Bilek [email protected]

206-AP: Phytochemical contents and antioxidant properties of early potatoes as affected by genotypes and cooking methods.

Reena Grittle PINHERO1, James A. SULLIVAN2, Qiang LIU3, Rong CAO3, Massimo MARCONE1, Rickey Y. YADA4

1 Department of Food Science, University of Guelph, Guelph, Ontario 2 Department of Plant Agriculture, University of Guelph, Guelph, Ontario 3 Agriculture and Agri-Food Canada, Guelph Food Research Centre, Guelph, Ontario 4 Faculty of Land and Food Systems, University of British Columbia, Vancouver Potatoes are a valuable dietary food due to their diverse functional ingredients including protein, fiber, vitamins and phytochemicals. Genotype, maturity, color of the flesh and skin and processing conditions can affect the content of phytochemicals. The objective of this study was to identify potato varieties, from eight varieties, with nutritional benefits with respect to their phytochemicals, antioxidant properties and the effect of cooking methods. Cooking methods included retrogradation (boiling followed by refrigeration at 4°C), retrogradation and reheating in the microwave, baking and microwaving. Phytochemicals such as total phenolics (TPC), total flavonoids (TFC) and total anthocyanins (TAC) contents were determined. Red Thumb had the significantly highest TPC in all processing methods followed by Purple Fiesta and Smart. Unlike TPC, the highest TFC was obtained in Purple Fiesta followed by Red Thumb, Smart and French Fingerlings. Anthocyanin contents were detected only in purple/red colored varieties such as Purple Fiesta, French Fingerlings, Ciklamen and Red Thumb. Purple Fiesta had the significantly highest TAC value followed by Red Thumb. Antioxidant properties were determined using three assays, ORAC, FRAP and DPPH. Significant differences were observed among varieties in their antioxidant properties. Highest DPPH activity was obtained in Purple Fiesta followed by Red Thumb, Smart, Adora and French Fingerlings. Highest FARP and ORAC values were obtained in Purple Fiesta and Red Thumb respectively. Based on phytochemical contents and antioxidant properties, Purple Fiesta, Red Thumb and Smart were identified as nutritionally superior varieties. Reena Grittle Pinhero [email protected]

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207-AP: Breeding the Boreal Series of Haskap (Lonicera caerulea) Robert .H. BORS 1 1. Department of Plant Sciences, University of Saskatchewan, SK The boreal series of haskap varieties was created with a strategy to combine desirable attributes of germplasm from Russia, Japan and the Kuril islands. Selection for vigour and general health occurred in the greenhouse. Field evaluations 5 to 7 years later were made of thousands of seedlings of which 200 superior selections were analyzed for fruit qualities in the lab. Breeding with diverse genotypes resulted in many selections exceeding parental germplasm for various desirable characteristics. For example, average fruit weights of Boreal Blizzard and Boreal Beauty are greater than other cultivars and about twice as heavy as most cultivars. Boreal Blizzard has half the acidity of most Haskap varieties while Boreal Beast is the only cultivar that ripens in August in Saskatchewan. Boreal Beast was chosen as a pollinizer for the other two Boreal varieties as it had compatible pollen and bloom time largely overlapped. Its flavour ratings were among the highest over several years of evaluation. The strategy of intercrossing diverse germplasm is resulting in transgressive segregation from which breakthroughs in haskap breeding are occurring. [email protected]

208-AP: Small fruit improvement program using biotechnology combined with conventional methods Samir C. DEBNATH* St John’s Research and Development Centre, Agriculture and Agri-Food Canada, Bldg. 25, 308 Brookfield Road, St. John’s, NL A1E 0B2, Canada Commercially important small fruit crop genera include but are not limited to Fragaria (strawberry; Rosaceae), Rubus (brambles: raspberry and blackberry; Rosaceae) and Vaccinium (blueberry, cranberry and lingonberry; Ericaceae). Lowbush blueberry (Vaccinium spp. L.), cranberry (V. macrocarpon Ait.) and lingonberry (V. vitis-idaea. L.) are three small fruit crops important to cool climates of Canada. These fruits are believed to have significant role in anti-tumor, anti-ulcer, anti-oxidant and anti-inflammatory activities. Wildberry production systems are changing to a more intensive cultivated system leading to an urgent need for developing new techniques for selecting and establishing high-yielding, pest tolerant small fruit crops which are well-adapted to diverse biotic and abiotic conditions. The presentation focuses on: wild berry germplasm collection, characterization, maintenance and their utilization in hybrid development. Bioreactor micropropagation in a liquid medium has been developed and found very successful for small fruit mass propagation. The total phenolic and oxygen radical absorbance capacity (ORAC) of blueberry tissue culture plants and of wild clones and cultivars of blueberries and cranberries were estimated. Hybrids obtained through crossing between half-high/highbush and lowbush blueberry genotypes and between Canadian and European lingonberries are being evaluated for frost, drought and pest resistance under greenhouse and field conditions. Biodiversity analysis in wild small fruit germolasm contributed significantly in planning future breeding program. In a field trial with cranberry wild clones and cultivars, effects of organic and inorganic fertilizers are being studied for quality fruit production and adaptability. Samir Debnath [email protected]

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209-AP: SNP molecular markers for the differentiation of specific ecotypes of the medicinal plant, Cudrania tricuspidata Bureau using ARMS-PCR and HRM curve pattern analyses Soo Jin Lee1, Yun-Hee Kim2, Shin-Woo Lee1* 1.Dept. of Agronomy & Medicinal Plant Resources, Gyeongnam National University of Science & Technology, JinJu, Republic of Korea 2. Dept. of Biology Education, College of Education, and IALS, Gyeongsang National University, JinJu, Republic of Korea Cudrania tricuspidata Bureau is one of the important Chinese medicinal perennial woody plants. Although C. tricuspidata is an important medicinal plant species registered in South Korea, no molecular markers are currently available to distinguish Korean-specific ecotypes from other ecotypes of different countries. In this study, based on single nucleotide polymorphisms among twelve individual ecotypes collected from different regions of South Korea and China, we were able to conclude that there are two different ecotypes in China and one in South Korea. By using specific molecular markers designed from SNPs derived from chloroplast MatK or TrnL-TrnF intergenic region and nuclear ribosomal DNA internal transcribed spacer (ITS) regions it was able to discriminate these three ecotypes via amplification refractory mutation system (ARMS)-PCR. In addition, high resolution melting (HRM) curve pattern analyses were also adopted for the identification of these distinct ecotypes. Our preliminary results indicate that further investigation must be followed up to look into the genetic diversity along with breeding program of such an important Chinese medicinal plant, C. tricuspidata. SHINWOO LEE [email protected]

210-AP: Deficit Irrigation Strategies for Improving the Aromatic Contents and Ripening in Gewürztraminer Grapes Yevgen KOVALENKO1*, Simone CASTELLARIN1 1. Wine Research Centre, University of British Columbia, Vancouver, BC Deficit irrigation of grapes is a viticultural practice is often used in red grape varieties to improve grape and wine quality; however, little work has been done with white grapes particularly in the Okanagan Valley. The timing of deficit irrigation application is also known to alter the effectiveness of the treatment. In this study, field-grown Gewürztraminer under a deficit irrigation regime applied at different periods throughout berry development (well-watered, pre-ripening deficit, post-ripening deficit, and prolonged deficit) were characterized for their eco-physiological, biochemical, and molecular response to the treatments. The hypothesis is that application of deficit irrigation will induce the synthesis of quality-determining volatile compounds with pre-ripening and prolonged application of water deficit being most effective than a later application. Treatments were replicated on four plots arranged in a randomized block design sampling every 7-14 days starting three week after fruit set. Grapevine leaf water potential was monitored using the Scholander pressure chamber at ± 1 h from solar noon. The effect of these treatments on leaf water potential, leaf gas exchange, and plant vegetative growth, as well as berry sugar, acid, and volatile organic compound concentrations was analyzed. Eco-physiological parameters, sugar concentration, berry weight, and vine yield were significantly affected by the deficit irrigation treatments, however, there was a clear effect from timing of application as well. At harvest, the free volatile profile was significantly different between treatments. The goal of this project is optimize grape quality through targeted practices as volatile compounds are closely tied to the economic potential of wine grapes. Yevgen Kovalenko [email protected]

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Biochemistry Posters (211-‐BC to 214-‐BC) Thursday July 6, FSC Atrium 11:20am -12:30pm 211-BC: Monitoring photosynthetic phenology using optically derived vegetation indices at the leaf-scale in temperate evergreen and deciduous forests. Christopher Y.S. Wong1,2* Altaf Arain3 & Ingo Ensminger1,2,4 1. Department of Biology, University of Toronto Mississauga, Mississauga, ON, Canada 2. Graduate Program in Ecology and Evolutionary Biology, University of Toronto, Toronto, ON, Canada 3. School of Geography & Earth Sciences, McMaster University, Hamilton, Ontario, Canada 4. Graduate Program in Cells and Systems Biology, University of Toronto, Toronto, ON, Canada Optically derived vegetation indices have been developed to provide information about plant status including photosynthetic activity. They can reflect changes in leaf pigments associated with light harvesting and dissipation and thus act as proxies of photosynthetic activity. In northern forests, which consist of deciduous and evergreen trees, there have been issues resolving evergreen phenology using the standard greenness index, the normalized difference vegetation index (NDVI). NDVI works well in deciduous trees, which exhibit a “visible” phenological process of leaf growth in the spring, and leaf senescence and abscission in the autumn. Evergreen conifers stay green year-round by investing structurally in their needles to retain them for multiple years and this involves “invisible” physiological changes within the needles that NDVI cannot resolve, so pigment sensitive vegetation indices have been suggested for evergreens. The aim of this study was to compare the photosynthetic phenology of evergreen and deciduous trees and the ability of vegetation indices to monitor them. We evaluated the greenness index, NDVI, and pigment sensitive indices: photochemical reflectance index (PRI) and chlorophyll/carotenoid index (CCI) in red maple, white oak and eastern white pine for two years. We also measured leaf gas exchange, chlorophyll fluorescence and photosynthetic pigment concentrations. We found that NDVI correlated with photosynthetic activity only in deciduous trees, whereas PRI and CCI correlated with photosynthesis across both evergreen and deciduous trees. These findings have implications to improve our use and understanding of remotely sensed vegetation indices as proxies of photosynthetic activity in northern forests for long-term monitoring. Christopher Wong [email protected] 212-BC: Designer lignin: Identification of p-hydroxybenzoyl-CoA monolignol transferase in poplar Heather A. MACKAY1, Yaseen MOTTIAR1, and Shawn D. MANSFIELD1 1. Department of Wood Science, University of British Columbia, 4030-2424 Main Mall, Vancouver, BC, Canada V6T 1Z4 Woody biomass utilization for the production of pulp and paper, and cellulosic biofuels requires significant investment of chemicals and energy. Lignin, a polyphenolic constituent of the secondary cell walls of vascular plants and the second most abundant biopolymer on Earth, is a significant impediment for wood processing. Delignification is critical for biomass utilization, and typically relies on the addition of acid or alkali to remove lignin for fibre separation in pulp and paper production, and to facilitate the saccharification of lignocellulosic material for biofuel production. To economize biomass utilization, researchers have focused on engineering lignin by manipulating the lignin biosynthetic pathway. Increasing the degree of acylation (the process of adding to an acyl group, or RCO– group, to a compound) of lignin monomers and/or integrating ester bonds into lignin polymeric backbone replacing the native ether bonds, increases the processability of the otherwise chemically-stable polymer to digestion upon alkali treatment. In Salix, Populus, and Palmae species, p-hydroxybenzoic acid (pHBA) acylate the monolignols as pendant groups via ester linkages,

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and as such increase the number of easily cleavable bonds available for degradation by mild chemical treatment. As a result, pHBA is of particular interest in the pursuit of designer lignin. p-hydroxybenzoyl-CoA monolignol transferase (pHBMT) is the putative enzyme thought to be responsible for decorating monolignols with pHBA, however, the gene encoding pHBMT remains elusive. Identifying pHBMT will allow us to better understand the mechanisms driving acylated monolignol biosynthesis and potentially develop novel transgenic plant varieties with unique lignin properties and commercial applications. Heather A. MacKay [email protected]

213-BC: Plant Cuticle Formation: TaFAR1 Contributes to Wax Biosynthesis in Bread Wheat Yulin Sun1*, Daniela Hegebarth1, Yong Wang2, Zhonghua Wang2, Reinhard Jetter1,3 1. Department of Botany, University of British Columbia, Vancouver, Canada 2. State Key Laboratory of Crop Stress Biology for Arid Areas, College of Agronomy, Northwest A&F University, Yangling, China 3. Department of Chemistry, University of British Columbia, Vancouver, Canada Bread Wheat (Triticum aestivum), one of the main crops in the world, provides roughly 20% of calories consumed by humans. The above-ground wheat surfaces are coated with cuticular wax, a natural water retention barrier consisting of very-long-chain fatty acids and their derivatives. However, the mechanisms of cuticular wax biosynthesis which lead to the complex chemical composition of wheat waxes remain largely unknown to date. In order to characterize biochemical pathways involved in the synthesis in primary alcohols, as one of the major components of wheat wax, a candidate gene was isolated from wheat cultivar Ming988 based on sequence similarity with Arabidopsis fatty acyl-CoA reductases generating wax alcohols and gene expression patterns. Heterologous expression of this gene, TaFAR1, in yeast (Saccharomyces cerevisiae) and Arabidopsis (Arabidopsis thaliana) confirmed its function in primary alcohol formation with substrate preference for very-long-chain fatty acyl-CoAs. The expression of TaFAR1 was highly induced by stress in wheat seedlings in an ABA-dependent manner. Drought treatment enhanced the expression of TaFAR1 and led to elevated accumulation of primary alcohols in wheat cuticular wax. Our results suggest that TaFAR1 plays a crucial role in wax primary alcohol biosynthesis and, thus, a key role in wheat cuticle formation.

Yulin Sun [email protected]

214-BC: Functional diversity among protease inhibitory cystatins in the plant kingdom Jonathan TREMBLAY1, Marie-claire GOULET1, Vanessa MERCURE GODIN, Charles GOULET1 and Dominique MICHAUD1 1 Centre de recherche et d’innovation sur les végétaux, Université Laval, Québec QC, Canada Numerous studies have discussed the potential of plant cystatins to implement resistance to coleopteran and acarian pests in transgenic crops. Once ingested, these proteins behave as competitive pseudosubstrate inhibitors in the herbivore, to block the active site of digestive proteases in the midgut lumen and compromise leaf protein digestion. To take full advantage of plant cystatins in pest control, a challenging task at present is to optimize their inhibitory profile towards arthropod proteases without altering host plant’s own proteases. In biochemical terms, the challenge is to identify or develop cystatin variants with both high affinity for the pest midgut proteases and low affinity for the host plant leaf proteases. We here performed protease inhibition assays with E. coli-produced forms of 20 phylogenetically distant plant cystatins and the proteases of selected plant and herbivorous arthropod species, to appreciate the extent of functional diversity among cystatins in the plant kingdom. Our results show a diversity of protease inhibitory profiles among currently known cystatins, and the usefulness of these proteins as a reservoir of protein templates for cystatin molecular improvement. We now apply a Loop Replacement Design approach to the

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model cystatins, with the aim of producing cystatin hybrids with distinct preferences for pest and host plant proteases. Jonathan Tremblay: [email protected]

Biotechnology/Tech Innovations Posters (215-‐BT to 218-‐BT) Thursday July 6, FSC Atrium 11:20am -12:30pm 215-BT: Recombinant protein accumulation patterns in the model expression host Nicotiana benthamiana Anne-Marie MALTAIS1*, Vanessa MERCURE-GODIN1, Marie-Claire GOULET1, Ann-Catherine LALIBERTÉ1, Charles GOULET1 and Dominique MICHAUD1 1. Centre de recherche et d’innovation sur les végétaux, Université Laval, Québec QC, Canada A correct assessment of recombinant protein yields in plant protein biofactories requires a careful monitoring of protein accumulation patterns at the whole-plant scale. We here monitored the accumulation patterns of four recombinant proteins transiently expressed in the widely used expression host Nicotiana benthamiana. Research efforts have been made in recent years to optimize environmental factors and cultural practices for this plant in a biopharmaceuticals production context. Little is still known, on the other hand, about how structural complexity and cellular destination of the expressed protein influence accumulation patterns in planta. We here used GFP variant pHluorin, protease inhibitor α1-antichymotrypsin, monoclonal antibody C5-1 and Influenza virus antigen hemagglutinin 1 as protein models of varying complexity to address this question. Variants of these proteins targeted to different cellular locations were expressed for a week in agroinfiltrated N. benthamiana leaves, and their abundance then evaluated in 11 sections of the plant encompassing old, mature and young leaves distributed on the main or axillary stems. Our data show variation in net yields from one protein to another, but a similar accumulation pattern for the four proteins, regardless of their final destination in the cell. These observations suggest that leaf position on the host plant has more impact on recombinant protein accumulation patterns than the origin, complexity or cellular destination of the protein expressed. Anne-Marie Maltais [email protected]

216-BT: Micropropagation and characterization of bigleaf maples (Acer macrophyllum) with valuable figured wood ZHOU, C., MATTSSON, J. Department of Biological Sciences, Simon Fraser University Some bigleaf maple (Acer macrophyllum) trees develop defects during wood formation that result in attractive ‘wavy figure’ patterns in the wood, which fetches a high price and is used primarily for making musical instrument and decorative products. While the underlying cause of these defects is unknown, the large extent and homogeneity of the defects in many individuals suggest an underlying genetic basis. Typically, available living material from cut trees is limited. To propagate from limited amount of source material, we have developed a procedure combining breaking apical dominance by cutting of shoot apices with in vitro hormonal induction of shoot multiplication, followed by root induction to generate a protocol for micropropagation of bigleaf maple. Since it is not given that clones of figured wood trees will develop figure and since harvested trees typically do not show figure in their 10-15 first growth rings, we have also begun searching for potential early anatomical and/or chemical differences between young clones from wildtype and figured trees. We have found that the average wood microfibril angle in clones of three independent figured trees deviate from wildtype clones. Pyrolysis Molecular Beam Mass Spectroscopy analysis also suggest that the content of two unknown cell wall components, mz_69 and mz_298, differ between wildtype and figured tree wood samples. We are extending this analysis to recently identified figured trees. Taken together, we have generated evidence that bigleaf maple can be propagated in vitro and also identified potential markers predicting figured wood formation.

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Chen Zhou [email protected]

217-BT: Genetic improvement of Canadian Salix for biomass production Emily K. MURPHY1*, Raju Y. SOOLANYAKANHALLY2, Shawn D. MANSFIELD1 1. Department of Wood Science, Faculty of Forestry, University of British Columbia, Vancouver, BC 2. Indian Head Research Farm, Agriculture and Agri-Food Canada, Indian Head, SK Increasing global populations, limitations in arable land, and extensive release of CO2 into the atmosphere are, collectively, driving the search for bio-based alternatives to the petroleum-derived fuel and chemical products that currently underpin the international economy. The inherent variability in cell wall characteristics of native willow (Salix spp.) populations make this tree species an attractive source of low-cost, high-volume biomass. Vetrix species, a sub-genus of Salix, have piqued interest, since they can be grown for biomass production in short-rotation woody crop (SRWC) systems, where they are cut back after the first year of growth to promote vigorous regeneration, and consequently higher biomass yields. High biomass yields from SRWC systems have encouraged bioenergy producers, but work remains to bring willow biomass to the forefront as an alternative energy source. To-date, Canadian breeding efforts have been focused on native varieties in an effort to exploit inherent resistance to local biotic and abiotic stressors. In 2013, a preliminary study of the genetic diversity of a collection of 46 native populations identified significant variation between individuals and amongst two distinct regional groups (east vs. west). The results of this study provide precedence for the characterization of biomass composition and genetic controls in native willow. In order to facilitate a range-wide exploration of phenotypic variation, 34 populations spanning the latitudinal range 45.21o to 55o are currently being screened for wood chemical composition, cell wall carbohydrates, enzymatic saccharification efficiency and physical properties in an attempt to direct future breeding and deployment strategies for Canadian biomass generation. Emily Murphy [email protected]

218-BT: Tissue culture of Cannabis sativa – Evaluating the effects of genotype and growth regulator combinations on shoot growth and plantlet production Danielle C. P. COLLYER1, Zamir K. PUNJA1 1.Department of Biological Sciences, Simon Fraser University, Burnaby, Canada Cannabis sativa is a high valued medicinal plant belonging to the family Cannabaceae. The availability of a tissue culture method can provide the opportunity to propagate disease-free plants in large numbers. The objective of this study was to quantify the growth response (measured as height, number of auxillary buds, number of auxillary shoots and stem width) of 3 cannabis strains (genotypes) [Pennywise (PWE), Girl Scout Cookies (GSC), Jesus OG (JOG)] on Murashige & Skoog (MS) medium. Plants were initiated from apical meristems dissected from shoot tips and grown on full-strength MS containing Gamborg B5 vitamins, sucrose (20g/L), activated charcoal (1g/L) and phytagel (3 g/L) (MS-C) supplemented with different growth regulators. Addition of 1 µM thidiazuron (TDZ) and 0.5 µM napthaleneacetic acid (NAA) produced more shoots (90% frequency) than giberellic acid and 6-benzylaminopurine, or TDZ alone. Apical meristems developed into shoots after 6 weeks at 25 ±2oC and at 102 µmoles m-2 s-1 light intensity, which were transferred to the same medium for elongation. Four weeks later, the mean heights of the three strains were not significantly (p=0.05) different (2.83, 2.38 and 2.12 cm for GSC, PWE and JOG, respectively). JOG produced slightly more auxiliary buds/shoot at 2.13 than GSC and PWE at 1.36 and 1.45, but this difference was not significant (p=0.05). The mean number of auxiliary shoots at 0.5 (GSC), 0.4 (PWE) and 0.32 (JOG) were not significantly different. Elongated shoots of GSC were transferred onto rooting medium (MS-C containing 37 µM indole-3-butyric acid and 40 µM silver nitrate), resulting in 76% rooting. Rooted plantlets were successfully acclimatized in soil. Danielle Collyer [email protected]

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Biotic Interactions Posters (219-‐BI to 227-‐BI) Thursday July 6, FSC Atrium 11:20am -12:30pm 219-BI: Caterpillar Herbivory as a Readout for Systemic Defenses Induced by Rhizosphere Microbes Christina L. WIESMANN1*, Li XIAO2, Lori SHAPIRO3, Lucy O’SULLIVAN4, Sophie KHORASANI4, Jiatong HAN1, Naomi E. PIERCE4, Frederick M. AUSUBEL5, Juli CARILLO2, and Cara H. HANEY1,2 1. Department of Microbiology and Immunology, The University of British Columbia, Vancouver, Canada 2. Land and Food Systems, The University of British Columbia, Vancouver, Canada 3. Department of Organismic and Evolutionary Biology, Harvard University, Cambridge 4. Department of Molecular Biology, Massachusetts General Hospital, Boston, MA 5. Michael Smith Laboratories, The University of British Columbia, Vancouver, Canada Strains of beneficial rhizosphere bacteria including Pseudomonas fluorescens can induce systemic resistance (ISR) or susceptibility (ISS) to foliar pathogens such as Pseudomonas syringae. We found that like ISR, ISS depends on the jasmonic acid receptor COI1; however, unlike ISR, ISS is independent of the salicylic acid pathway NPR1 protein. There is a well-studied antagonistic interaction between the plant hormone salicylic acid (SA), which is involved in resistance to bacterial pathogens, and jasmonic acid (JA), which is involved in resistance to chewing insects. This antagonistic interaction predicts that P. fluorescens stains capable of inducing ISR to bacteria will confer susceptibility to insect herbivory while ISS strains will confer resistance to insects. We tested whether P. fluorescens has an effect on Arabidopsis herbivory by the cabbage looper Trichoplusia ni. We found that treatment of Arabidopsis roots by both P. fluorescens ISS and ISR strains resulted in increased resistance to caterpillars, suggesting that P. fluorescens is perturbing JA/SA antagonism. We found that while SID2 and NPR1 similarly increase resistance to caterpillars with no bacteria, resistance to caterpillars induced by all P. fluorescens strains depends on a common set of genes and is SID2 dependent and NPR1 independent. This suggests that all P. fluorescens strains tested have a single mechanism to induce susceptibility to P. syringae and resistance to T. ni, while only a subset of P. fluorescens strains have an independent mechanism to induce ISR. We are currently exploring the molecular mechanism by which P. fluorescens modulates JA/SA antagonism and promotes herbivore resistance. Christina Wiesmann [email protected] 220-BI: Characterizing root-associated fungal exudate profiles and their impacts on plant growth Joshua J.R. FRANK1*, Mamdouh ABOU-ZAID1,2, Tod RAMSFIELD3, Danielle WAY1,4 1. Department of Biology, Western University, London, ON 2. Great Lakes Forestry Centre, National Resources Canada, Ottawa, ON 3. Northern Forestry Centre, National Resources Canada, Edmonton, AL 4. Nicholas School of the Environment, Duke University, Durham, NC Root-associated fungi exude compounds into the rhizosphere that can promote plant growth, initiate fungal-plant symbioses, and reduce competition with other microorganisms. However, the chemical identity of these exudates and their impacts on plant growth are largely unknown. To address this, we examined fourteen fungal isolates, taken from aspen roots, to generate isolate-specific exudation profiles and determine the effects of root-associated fungal exudates on plant growth. We hypothesized that: 1) all isolates would exude organic acids and amino acids, but the exudate profile would vary between isolates; and 2) fungal exudates would promote plant growth. Fungal exudates were extracted from the liquid fungal growing medium and analyzed using HPLC-MS to develop an exudation profile of the fungal compounds. We found that isolates exude organic acids, vitamins, plant hormone derivatives,

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and amino acids. Furthermore, isolate-specific exudation profiles were observed. The effects of fungal exudates on plant performance were varied: Arabidopsis inoculated with fungal exudates from some isolates had up to a 150% reduction in mean rosette area and earlier flowering, while, other fungal exudates increased plant dry weight by up to 85%. As well, exudates from two fungal isolates increased seedling mortality, with one leading to 100% mortality of seedlings. Our results suggest that root-associated fungi exude a wide range of biologically active compounds in an isolate-specific manner, and that these exudates are not necessarily beneficial for plant growth. Furthermore, isolate-specific exudate profiles could be used to help identify and distinguish root-associated fungi from one another. Joshua Frank [email protected]

221-BI: Functional characterization of mycorrhiza responsive genes

Kishore VISHWANATHAN1,2*, Cara HANEY2, Andrea POLLE1

1. Department of Forest Botany and Tree Physiology, Georg-August-Universität Göttingen, Germany 2. Department of Microbiology and Immunology, University of British Columbia, Canada

Mycorrhization is a symbiotic association of plant roots and some fungi. It is mainly characterized by a continuous exchange of nutrients and carbon between the symbionts. Persistent communication exists between the partners during the course of symbiosis including pre-colonization, colonization and mature symbiosis. The genetic basis of symbiosis between ectomycorrhizal fungi and their tree hosts has remained largely intractable because of the lack of tools and long lifespan of their hosts. Arabidopsis thaliana is a model plant that is not colonized by mycorrhizal fungi. However, Arabidopsis responds to the ectomycorrhizal fungus Laccaria bicolor by increasing lateral root abundance in the pre-colonization phase. We hypothesized that Arabidopsis may contain some of the same genes required for a subset of host responses to mycorrhizal fungi. We tested well-characterized Arabidopsis genes involved in growth, development and defences for increased lateral root formation in the presence of L. bicolor. We identified several genes involved in defence that are required for response to mycorrhizal fungi in Arabidopsis. Though mycorrhization can influence various aspects of plant growth and development, it is also known to contribute to plant defence against pathogens and herbivores. We found that when challenged with the plant pathogen Pseudomonas syringae pv. tomato, L. bicolor primed Arabidopsis plants for systemic defences. To identify the genetic basis of mycorrhizae defence priming, we are testing genes we identified as required for mycorrhizae-induced increased lateral root formation and known genes required for induced systemic resistance against other pathogens. Long term, we hope to use Arabidopsis to identify genes that may be involved in the early colonization of mycorrhization or modulation of systemic defences.

Kishore Vishwanathan [email protected]

222-BI: Use of Pseudomonas fluorescens FW300-N2C3 in competition assay to screen for plant growth-promoting bacteria

Sarzana HOSSAIN1,, Ryan MELNYK1, and Cara HANEY1 1. Department of Microbiology and Immunology, University of British Columbia, Vancouver, BC The microbiome of the plant rhizosphere harbours diverse phyla of bacteria, where many members of this environment act as plant growth-promoting (PGP) bacteria. One avenue by which PGP bacteria benefit their host plant is by preventing or compromising growth of phytopathogenic bacteria by out-competing themway of competition over them. We have developed a competition assay for Arabidopsis thaliana, from which we can easily identify strains that are competitively dominant and protect against the opportunistically pathogenic Pseudomonas fluorescens FW300-N2C3 (N2C3). When the roots of sterile A. thaliana seedlings are inoculated with N2C3 is used to inoculate the roots of sterile A. thaliana seedlings, it effectively colonizes the root, impairs plant growth and ultimately leads to death. However, co-inoculation with certain PGP strains can restore normal plant growth. We have used this observation to develop an assay that assessesto measure a strain’s potential to protect seedlings from N2C3 as measured by seedling survival. .This in vivo assay provides a simple assessment to determine if a strain protects or not, as the result is on the basis of the A. thaliana plants surviving or succumbing to N2C3 infection. The assay does not evaluate directly determine if the success of a rescuing strain is due to direct antagonism of N2C3, outcompeting N2C3, growth-promoting interactions with the host plant, or a combination of these effects. We have

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performed this assay with 13 Pseudomonas and related strains, where 5 rescued normal plant growth and 8 did not. Future experiments will be focused on understanding the mechanism by which PGP can protect plants from opportunistic pathogens.

Sarzana Hossain [email protected]

223-BI: Cultivable bacteria associated with the giant kelp Macrocystis pyrifera can degrade and metabolise alginate Laura W. PARFREY1,2, Jordan LIN1 1. Department of Botany, University of British Columbia, Vancouver, BC 2. Department of Zoology, University of British Columbia, Vancouver, BC Bacteria are a critical component of marine carbon cycling. They transfer organic carbon to higher trophic levels and remineralise it into inorganic forms. Within the coastal environment, kelp forests are highly productive ecosystems and a large source of organic carbon. Bacterial cycling of kelp carbon is poorly understood, despite the magnitude of biomass kelp represent. We surveyed the uncultured and cultivable communities of the giant kelp Macrocystis pyrifera, a foundational species in Pacific kelp forests. We also assayed bacterial isolates for the degradation and metabolism of alginate, the primary polysaccharide in kelp tissue. A large portion of cultivable bacteria (66%) demonstrated some type of alginate activity, yet cultured taxa that interacted with alginate were poorly represented in uncultured communities. Alginate degradation and metabolism are not phylogenetically restricted but instead found across diverse genera in the Bacteroidetes and Proteobacteria. Alginate activity also varied between isolates, even those with identical 16S genes, thus taxonomy may be a poor indicator of function. The broad distribution of alginate activity and its poor phylogenetic signal in cultivable bacteria suggests that it may be widespread. Bacterial interaction with alginate may thus be a redundant trait in kelp forests and could represent an important driver in community assembly. Jordan Lin [email protected]

224-BI: Buckwheat as a pre-plant soil amendment provides control of seedling damping-off and root rot of radish and cucumber Pervaiz A. ABBASI, Willy Renderos Agriculture and Agri-Food Canada, Kentville Research and Development Centre, Kentville, NS Buckwheat is a fast-growing cover crop with a potential to improve soil quality and health. In this study, the impact of buckwheat plant material (BPM) as a soil amendment on plant growth and suppression of seedling damping-off and root rot of radish and cucumber caused by Rhizoctonia solani or Pythium ultimum, respectively was investigated in pot assays. Fresh BPM grown in a greenhouse potting mix for 4-5 weeks was chopped and incorporated at various rates into a pathogen (R. solani or P. ultimum inoculum produced on sterilized rye seed) infested field soil. The soil was then incubated for various time periods prior to planting radish or cucumber seeds, and the effects of BPM on plant growth and disease development were determined 2 weeks after planting. There was no disease protection if radish or cucumber seeds were planted within 2 weeks after incorporating BPM in the infested soil. Disease control was evident when planting was delayed for 3 weeks after amending BPM in the infested soil. The BPM treatments provided modest to good control of radish plants from Rhizoctonia damping-off (41-66% healthy and 2.2 mean severity compared to 1-9% healthy and 3.4-4.2 mean severity in the control) and good control of cucumber plants from Pythium damping-off and root rot (81% healthy and 1.7 mean severity compared to 22% healthy and 3.4 mean severity in the control). The effect of BPM amendment on plant dry weights, soil pH, and soil microbial populations of indigenous bacteria and fungi is also discussed. [email protected]

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225-BI: Characterization of Pseudomonas fluorescens mutants impaired for Arabidopsis thaliana rhizosphere colonization Zhexian Liu1,2, Lori Shapiro3, Sophie Khorasani4, Cara Haney1,2 1. Department of Microbiology and Immunology, University of British Columbia, Vancouver, BC 2. Michael Smith Laboratories, University of British Columbia, Vancouver, BC 3. Department of Organismal and Evolutionary Biology, Harvard University 4. Department of Molecular Biology, Massachusetts General Hospital Pseudomonas fluorescens, an important constituent of the plant root microbiome, contributes to plant health by promoting plant defenses and facilitating nutrient uptake. Though the beneficial effects of P. fluorescens are well-documented, the mechanisms through which P. fluorescens colonizes plant roots and induces plant phenotypic changes are poorly understood. To facilitate elucidating the molecular mechanisms of host colonization, we sequenced the genome of P. fluorescens strain WCS365, a particularly successful plant rhizosphere colonizer. We then performed a screen using transposon mutagenesis and high-throughput sequencing (Tn-Seq), which identified 93 candidate genes potentially implicated in root colonization of either wild-type or immunocompromised Arabidopsis thaliana. We have selected a subset of bacterial genes identified in the TnSeq screen for further characterization. We have generated clean deletions of these candidate genes and are currently characterizing the in vitro and in planta phenotypes of the mutants, including biofilm formation, swimming motility, and surface motility. From the initial Tn-seq screen, a number of transposon mutants were incapable of colonizing wild-type A. thaliana, yet were unimpaired in the colonization of an immunocompromised host. These observations suggest that these genes may be responsible for plant immune evasion in P. fluorescens WCS365. Further characterization of this candidate gene may reveal the role of motility and biofilm formation in plant rhizosphere colonization. Zhexian Liu [email protected]

Technological Innovations Posters (226-‐BI to 227-‐BI) Thursday July 6, FSC Atrium 11:20am -12:30pm Note: these two posters are clustered with Biotic Interactions but are thematically distinct. 226-BI: High Resolution Melt Analysis as an SNP Genotyping Tool David Bird1, Jonathan Roveredo1 1. Department of Biology, Mount Royal University, Calgary Canada Single nucleotide polymorphisms (SNPs) are a common type of mutation, and are often used in model systems in plant biology. These SNPs are typically sequenced following amplification of the region of interest by the polymerase chain reaction (PCR). Sequencing this way is done in order to track the SNP through generations, or through any crossing that is done in order to further isolate the desired SNP, or to distribute the SNP of interest into a desired genetic background. However, using sequencing to verify the presence of an SNP can be expensive and time consuming. High resolution melt analysis (HRM) is a technique used to precisely measure the annealing temperatures of the product DNA during quantitative PCR. Here, we report HRM analysis to screen for tt4-4, an SNP allele of the TRANSPARENT TESTA4 (TT4, At5g 13930) gene in A. thaliana. With HRM we can accurately determine the presence and zygosity of the tt4-4 allele in each plant based on the slight differences in annealing temperatures between the amplicons produced by the wildtype and mutant alleles. HRM provides us with a fast and inexpensive method of genotyping SNPs, in comparison with sequencing. Jonathan Roveredo: [email protected]

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227-BI: Developing targeted gene editing in a wheat mesophyll protoplast system using CRISPR/Cas9 Xiucheng CUI1,2, Gopal SUBRAMANIAM1, Thérèse OUELLET1 1Ottawa Research and Development Centre, Agriculture and AgriFood Canada, Ottawa, ON 2Department of Biology, University of Ottawa, Ottawa, ON The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system has become a promising tool for targeted gene editing in a variety of organisms including plants. In this system, a 20nt sequence on a single guide RNA (sgRNA) is the only gene-specific information required to modify a target gene. In this project, we are using the CRISPR/Cas9 system to modify three wheat genes identified in previous experiments, including an ABC transporter, the Nuclear Transcription Factor X box-binding-Like 1 (NFXL1), and a non-specific Lipid Transfer Protein (nsLTP). Two sgRNAs were designed for each gene and were shown in an in vitro CRISPR/Cas9 assay to guide the sequence-specific cleavage of the DNA template with high efficiency. An in vivo assay for CRISPR/crCas9 was established by optimization of a wheat mesophyll protoplast isolation and transformation system. Using a GFP construct as a positive control, estimated transformation efficiencies of about 60% are routinely obtained. High throughput sequencing of PCR amplicons including the sequences targeted for gene editing by CRISPR/Cas9 has clearly showed that the three targeted genes have been successfully edited in the protoplast system with efficiencies up to 42%. A comparison of editing efficiencies in wheat protoplasts between the crCAS9 and the pcoCAS9 will also be presented. [email protected]

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Cell Wall Posters (228-‐CW to 235-‐CW) Thursday July 6, FSC Atrium 11:20am -12:30pm

228-CW: Involvement of reactive oxygen species in spore wall development in the moss, Physcomitrella patens Fazle RABBI1*, Neil W. ASHTON2, Dae-Yeon SUH1 1. Department of Chemistry and Biochemistry, University of Regina, Regina, SK 2. Department of Biology, University of Regina, Regina, SK Reactive oxygen species (ROS) function in a tightly regulated manner and play important roles in a wide range of cellular development processes and defense mechanisms in plants. The level of ROS is regulated by various ROS production and scavenging enzymes. The main constituent of the exine and perine layers of Physcomitrella patens spore wall is sporopollenin. We hypothesized that ROS are involved in formation of the spore wall of P. patens. To evaluate the roles of ROS in spore wall formation, we exposed sporophytes containing developing spores to three different ROS scavengers, namely ascorbic acid, dimethylthiourea and 4-hydroxy-TEMPO. This resulted in the development of spores with weakened spore wall as well as spores without a perine layer. The former condition was detected by studying osmolysis of the spores in the presence of distilled water. Our results suggest that ROS are involved in moss spore wall formation and also, therefore, possibly in the formation of sporopollenin. Fazle Rabbi [email protected]

229-CW: The Class II KNOX genes KNAT3 and KNAT7 work cooperatively to activate syringyl lignin biosynthesis and regulate secondary cell wall deposition in Arabidopsis

Shumin Wang1*, Masatoshi Yamaguchi2, Etienne Grienenberger3, A. Lacey Samuels1, Shawn D. Mansfield4 and Carl J. Douglas1 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada 2. Graduate School of Science and Engineering, Saitama University, Saitama, Japan 3. Institut de biologie moléculaire des plantes (IPMB), CNRS UPR 2357, Strasbourg, France 4. Department of Wood Science, University of British Columbia, Vancouver, BC, Canada KNOTTED1-like homeobox (KNOX) genes are evolutionarily ancient. Gene duplication events and diversification of KNOX genes have produced two classes, Class I (KNOX1) and Class II (KNOX2) in land plants. In Arabidopsis, KNAT7, one of the four Class II KNOX genes, acts as a negative regulator of secondary cell wall biosynthesis in interfascicular fibres, but knat7 loss of function mutants have an irregular xylem (irx) phenotype, suggesting a potential positive regulatory role in xylem vessel secondary cell wall deposition. However, the functions of other three KNOX2 genes, KNAT3, KNAT4 and KNAT5, in secondary wall formation still remain unclear. We found that Arabidopsis knat3 knat7 double mutants had altered stem biomechanics with decreased tensile strength and tensile modulus of elasticity compared with wild type and single mutants. Also, knat3 knat7 loss-of-function mutants exhibited enhanced irx phenotypes relative to the single mutants, and decreased fiber cell wall thickness. Lignin content analysis showed that knat7 had increased lignin content, while knat3 knat7 double mutants had no change in lignin content. The transcript abundance of FERULATE-5-HYDROXYLASE (F5H), which encodes a key enzyme in the biosynthesis of S lignin, was strongly decreased in the knat3 knat7 mutants, but was little affected in the single mutants. Consequently, the S to G ratio was extremely reduced in the knat3 knat7 mutants relative to wild-type plants. Our study suggests that in addition to KNAT7, other KNOX2 genes also contribute to plant stem strength by affecting the cell wall deposition and the integrity of cell-wall matrix, and that a complex network of positive and negative regulation by KNOX2 proteins regulates secondary wall formation. [email protected]

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230-CW: Using functional genomics to discover novel secondary cell wall genes involved in lignin content variation in poplar Lan T. TRAN1*, Jürgen EHLTING2, Carl J. DOUGLAS1 1. Department of Botany, University of British Columbia, Vancouver, BC 2. Centre for Forest Biology and Department of Biology, University of Victoria, Victoria, BC Secondary cell walls (SCWs) contain a significant amount of fixed carbon that can be harnessed to produce renewable energy. However, efficient conversion of wood-based biomass for use as an alternative fuel source is hampered by lignin. Many aspects of SCW biosynthesis remain unknown including how genes of broad functional classes affect lignin deposition. A large-scale genetic association (GA) mapping study in poplar (Populus trichocarpa) previously identified 25 novel genes associated with lignin content and composition variation. Using this gene set, we are testing the hypothesis that these genes influence lignin content and/or composition by characterizing the biological functions for a subset of these genes. Here Solute Carrier 15 (SLC15), a member of the peptide/nitrate transporter superfamily is described. SLC15 is expressed throughout the plant in the vascular tissue. Transgenic poplar SLC15 RNA-interference (RNAi) lines showed decreased phenolic content compared to wild-type. Under high exogenous nitrogen conditions, SLC15-RNAi lines produced wood with decreased syringyl lignin compared to wild-type. Together, these findings suggest that SLC15 could help regulate nitrogen allocation throughout the plant. Understanding how previously unsuspected classes of genes identified through unbiased approaches such as GA mapping will contribute to our understanding of wood development and will enable the optimization of poplar for a diversity of applications. Lan Tran [email protected] 231-CW: Using Brachypodium distachyon as Model Species to Study the Secondary Cell Wall Synthesis in Monocots Xuan YANG1,2*, Kathleen HILL1, Lining TIAN2

1 Department of Biology, University of Western Ontario, London, Ontario, Canada 2 London Research and Development Centre, Agriculture and Agri-Food Canada, London, Ontario, Canada Cell wall performs several essential functions in plants, such as structural support, interface of intercellular communication and interaction between plants and their environment. Since secondary cell wall makes up the most of the vegetative biomass of mature plants, cellulose synthase (CESA) genes that are involved in secondary cell wall formation (CESA4, CESA7 and CESA8) are the major targets of this research. To study the cellulose synthesis in secondary cell wall and its potential regulatory network in monocot plants, the expression patterns of CESA genes were examined in Brachypodium distachyon throughout different developmental stages. At most developmental stages, BdCESAs are highly expressed in stems compared to other organs. BdCESA4 and BdCESA7 showed a similar expression pattern across different stages, while BdCESA8 displayed a somewhat different expression pattern from BdCESA4 and BdCESA7. In general, the expression levels of BdCESA4 and BdCESA7 are slightly higher in leaves than in roots, but remained a lower expression level compared to stems. At the early developmental stages, the lowest expression of BdCESA8 was found in leaves. However, BdCESA8 expression level in leaves increased rapidly at the later developmental stages and reached its peak at the fruit development stage. This peak is also higher than BdCESA8 expression in both roots and stems at the same stage. The different expression patterns of BdCESA8 suggest that this gene plays a different role in secondary cell wall synthesis from BdCESA4 and BdCESA7.

Xuan Yang [email protected]

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232-CW: Expression of bacterial chorismate pyruvate lyase in poplar leads to an increase in p-hydroxybenzoylated lignin Yaseen MOTTIAR, Shawn D. MANSFIELD Department of Wood Science, University of British Columbia, 2424 Main Mall, Vancouver, BC The lignin of poplar and willow trees is notable for the presence of p-hydroxybenzoyl moieties (–pHB) that occur as ester-linked pendant groups. Grasses analogously incorporate p-coumaroyl groups that become acylated to lignin by specialized monolignol acyltransferase enzymes. Ongoing work is aimed at identifying the poplar acyltransferase(s) responsible for p-hydroxybenzoylated lignin. The substrate of this enzyme – p-hydroxybenzoic acid - is likely produced in poplar from p-coumaric acid by way of an unidentified biosynthetic mechanism. In contrast, bacteria possess a chorismate pyruvate lyase (CPL) enzyme that generates p-hydroxybenzoic acid directly from chorismate, a key shikimate pathway intermediate and lignin precursor. Expression of bacterial CPL in poplar led to hyperaccumulation of p-hydroxybenzoic acid glycosides as well as increased levels of –pHB groups decorating the lignin. These plants also seem to be deficient in lignin owing to a decreased carbon flux from chorismate into the phenylpropanoid pathway. These results illustrate how the inherent metabolic plasticity of lignin biosynthesis can be harnessed to produce designer lignins with novel physiochemical properties. [email protected]

233-CW: Identification of a seed coat-specific promoter fragment from the Arabidopsis MUCILAGE-MODIFIED4 gene Gillian H. DEAN1, Zhaoqing JIN1, Lin SHI1, Elahe ESFANDIARI1, Robert MCGEE1, Kylie NABATA1, Tiffany LEE1, Ljerka KUNST1, Tamara L. WESTERN2, George W. HAUGHN1 1 Dept. of Botany, University of British Columbia, 6270 University Blvd., Vancouver, BC, V6T 1Z4, Canada. 2 Dept. of Biology, McGill University, 1205 Ave. Docteur Penfield, Montreal, QC, H3A 1B1, Canada. The epidermal cells of the Arabidopsis seed coat produce and secrete large quantities of mucilage during differentiation. In mature seeds, hydration causes the mucilage to be extruded to form a tightly-attached halo around the seed. As mucilage is composed mainly of pectin, and also contains the key cell wall components cellulose, hemicellulose, and proteins, it is a valuable model for studying numerous aspects of cell wall biology. Seed coat-specific promoters are an important tool that are required to assess the effects of expressing biosynthetic enzymes and diverse cell wall-modifying proteins on mucilage structure and function, and can also be used for production of easily accessible recombinant proteins of commercial interest. The MUCILAGE-MODIFIED4 (MUM4) gene is expressed in a wide variety of plant tissues and is strongly up-regulated in the seed coat during mucilage synthesis, implying that the promoter contains a region responsible for seed coat-specific expression. Promoter deletion analysis allowed the isolation of a 308 base pair sequence (MUM40.3Pro) that directs reporter gene expression in the seed coat cells of both Arabidopsis and Camelina sativa. MUM40.3Pro is regulated by the same transcription factor cascade as endogenous MUM4, and represents a promoter fragment can be used as a tool for seed coat biology research. Gillian Dean [email protected]

234-CW: Fasciclin-like arabinogalactan proteins 11, 12 (AtFLA11 and, AtFLA12) in are integral to early sSecondary cCell wWall synthesis Lynn CHEN and Shawn D. MANSFIELD

Department of Wood Science, University of British Columbia, 4030-2424 Main Mall, Vancouver, BC, Canada V6T 1Z4 In terrestrial plants, the process of cell wall biosynthesis is as fundamental and important as photosynthesis. The modulation of cell wall biosynthesis directly influences many aspects of plant growth and development including,

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but not limited to, cell division and expansion, plant morphogenesis, response to environmental cues, and ultimately the overall structure of the plant. A thorough understanding of the genes and gene networks regulating the biosynthetic processes governing the deposition polymers, such as cellulose is pivotal to understanding plant growth. Abstract: Plant development is regulated by a gene network. Gene manipulation is able to study plants traits for the interest of both bioengineering and botany, such as flower determination and secondary cell wall synthesis. TheIt has been shown that former studies has shown that the family of fasciclin-like arabinogalactan proteins (FLAs) are actingcontribute to the synthesis of in secondary cell wall cellulose deposition in Arabidopsis. By using Using an inducible master transcription factor controlling xylem cell fate, VASCULAR NAC-DOMAIN7 (VND7) fused to a glucocorticoid receptor (GR), living protoxylem cells were visualized by live-cell imaging. Fusing the two principal Taku Demura’s VND7 system, it’s available to observe FLA genes (AtFLA11 and AtFLA12) s activities fusing with a fluorescence reporter gene and their native promoter and also to seewe examined the interactions with other importantof the Fasciclin-like arabinogalactan proteins with proteins specific to secondary cell wall biosynthesis, such as cellulose related genes such as AtCesa7. By using cConfocal microscopy e we could confirmed that both AtFLA11 and AtFLA12 are indeed co-localized on to the secondary cell wall, . Aand when during active secondary cell wall cellulose deposition AtCesa7 involved complexes are synthesizing the cellulose microfibrils, AtFLA11 and AtFLA12 have been delivered to the appear to be localized outside of the plasma membrane which leads us to observesuggests that FLA are integral for the early stages of secondary cell wall synthesiss in future studies.

235-CW: A Novel XTH-related Endo-Glucanase from Vitis vinifera Degrades (1,3/1,4) Mixed-linkage Glucans with Unique Specificity Nicholas MCGREGOR1,2, Victor YIN1,2, Ching-Chieh TUNG3, Filip VAN PETEGEM3, Harry BRUMER1,2,3,4 1. Department of Chemistry, University of British Columbia, 2063 Main Mall, Vancouver, BC 2. Michael Smith Laboratories, University of British Columbia, 2185 East Mall, Vancouver, BC 3. Department of Biochemistry and Molecular Biology, University of British Columbia, 2350 Health Sciences Mall, Vancouver, BC 4. Department of Botany, University of British Columbia, 6270 University Boulevard, Vancouver, BC Xyloglucan is a major hemicellulose in the primary cell wall of all land plants. Enzymes from the xyloglucan endo-transglycosylase/hydrolase (XTH) subfamily of glycoside hydrolase family 16 (GH16) are unique in their ability to remodel xyloglucan in vivo and facilitate primary cell wall expansion. Phylogenetic analyses of available plant genome sequences have revealed a novel family of XTH-like enzymes in plants. The endo-glucanase GH16 (EG16) family represents an extant evolutionary intermediate between bacterial licheninases and plant XTHs. Structural and biochemical characterization of a representative EG16 from Vitis vinifera (common grape vine) reveals broad specificity towards β(1,4) glucans. Kinetic subsite mapping demonstrates the existence of an extended active site cleft catalyzing the hydrolysis of a variety of substrates with a preference for cereal mixed-linkage glucans. X-ray crystallography reveals a wide, yet linear, active site cleft which accommodates both xyloglucan, a branched cross-linking glucan, and mixed-linkage glucan, a linear β(1,4) cross-linking glucan with β(1,3) “kinks”. Analysis of hydrolysis products from different plant cell wall polysaccharides reveals that this enzyme catalyzes the hydrolysis of mixed-linkage glucans in a novel way. This unique specificity causes remarkable changes to mixed-linkage glucan, increasing its propensity to form gels. Although the biological function of EG16 orthologs remains unclear, expressed sequence tag data and genomic abundance, lead us to expect an important, as-yet-unknown biochemical role for these enzymes. Nicholas McGregor [email protected]

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Development Posters (236-‐DV to 248-‐DV) Thursday July 6, FSC Atrium 11:20am -12:30pm 236-DV: Defining Insulator Function in Plants Anh TRAN1*, Douglas A JOHNSON1 1. Department of Biology, University of Ottawa, Ottawa, Ontario, Canada The need to discover and analyze plant specific regulatory elements that can be used to control transgenic gene expression is necessary. Chromatin insulators are critical DNA elements capable of doing so by controlling unwanted interactions or misexpression naturally within the genome. Insulators block enhancer-promoter interactions as well as act as chromatin barriers. The functionality of insulators in gene regulation is thought to be dependent on primary sequence-specific DNA binding proteins as well as secondary or “bridging” proteins that create chromatin loops or boundaries restricting gene expression. Currently the most extensively analyzed insulators occur in non-plant species. This study aims to test two of these insulators, UASrpg from the fungus Ashbya gossypii and BEAD1c from human T-cell receptors to determine potential insulator function in the model plant Arabidopsis thaliana to understand better the models and mechanisms of plant insulators. Candidate insulators were analysed by creating inversions of the sequence as well as mutations in potential protein binding sites and testing for their activity on the seed specific Napin promoter controlling the expression of a GUS reporter gene. Results to date have shown that mutations in UASrpg and BEAD1c significantly reduce insulator activity suggesting that plant insulators may use similar DNA sequences and thus pathways found in other species. They also argue for multiple insulator sequences and conservation of insulator pathways across kingdoms. Anh Tran [email protected]

237-DV: Using green algae as models for the evolution of multicellularity Carmen MARQUEZ-MELLIDEZ1*, Kenneth WILSON1 1. Department of Biology, University of Saskatchewan, Saskatoon, SK Green algae (Chlorophyta) are used as model organisms for many biological processes, from photosynthesis to toxicology. In the study of the origin of plants, Chlorophyta and Streptophyta, the clade which contains the higher plants, share common ancestors. Their similarities at the can be linked back to the endosymbiotic origin of the chloroplast. Green algae and higher plants share similar photosynthetic proteins and complexes that are postulated to have evolved from cyanobacteria. However, the two groups exhibit convergent evolution of multicellularity. The most highly studied model system for examining the evolution of multicellularity in green algae is the order Volvocales. Examining the members of this order we can investigate the transition from single celled Chlamydomonas spp., to undifferentiated colonial genera such as Gonium, which has 4 to 16 cells, to Volvox a colonial organism made up of hundreds of cells. In Volvox we also see cellular differentiation, with somatic and germ cells. Important traits such as size, reproduction strategies and genetic information have been used to build a timeline to explain evolution within the Chlorophyta. Starting with unicellular Chlamydomonas, the evolution of the Volvocales will be discussed as a model to understand the development of complex forms of life. Carmen Marquez-Mellidez [email protected]

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238-DV: Evidence of seed-derived auxins coordinating fruit development in pea (Pisum sativum) Dilini Adihetty*, Jocelyn Ozga, and Dennis Reinecke Plant BioSystems Group, Department of Agricultural, Food, and Nutritional Science, University of Alberta, Edmonton, AB, Canada T6G 2P5 In pea (Pisum sativum), normal pericarp growth requires the presence of seeds and removal or abortion of seeds leads to reduced pericarp growth and subsequent abscission. Previous studies showed that auxin levels are higher in seeds than the surrounding ovary (pericarp) tissues, suggesting that seed-derived auxins are transported to the surrounding tissues to promote pericarp (ovary) growth and development. However, direct evidence for auxin transport from the seeds to the fruit is lacking. To address this, we have investigated the spatial pattern of auxin distribution in pericarps and associated fruit tissues using an auxin-inducible DR5::GUS reporter system in pea. We observed higher GUS staining and nearly two-fold greater GUS activity in 4 days after anthesis fruit (funiculus, ventral pericarp suture, and pericarp wall) and attachment (pedicel and peduncle) tissues when the fruit contained developing seeds compared to tissues from deseeded fruit. The polar auxin transport (PAT) inhibitor, 1-N-Naphthylphthalamic acid (NPA) applied to the peduncles of fruit with developing seeds increased GUS staining and activity in tissues above the application point. Application of the PAT inhibitor directly to the pericarp wall, of seeded fruits, also increased GUS staining and activity in the fruit tissues. However, application of NPA to either the peduncle or ovary of deseeded fruits did not affect GUS staining or activity of the fruit or attachment tissues. These data support that seeds act as a source of auxins for developing pea ovaries and that auxin transport from the seeds to the fruit and attachment tissues is at least partially mediated through polar transport pathways.

Dilini Adihetty; [email protected]

239-DV: SQUAMOSA Promoter Binding Protein-like (SPL) Genes in Barley and Their Role in Plant Development

Rajiv K. TRIPATHI1*, Jaswinder SINGH1

1. Plant Science Department, 21 111 Rue Lakeshore, McGill University, Quebec, H9X 3V9, Canada

Barley is one of the oldest, widely distributed and economically important crops. Diploid nature and high collinearity of its chromosomes with other Triticae crops makes barley a great model for genetically complex cereals such as wheat. Growth and development of barley is under strict molecular regulation which requires thorough investigation. Involvement of SQUAMOSA-promoter binding like (SPL) gene family has been described for their possible role in the transition of vegetative to reproductive phase in other plant species. However, knowledge about this gene family in barley is limited. To investigate the regulation of these phenomena in barley, we studied the interaction between SPL, miR156 and miR172 regulatory hubs. We found that barley genome contained 13 SPL gene family members which were widely distributed on different chromosomes. We identified that 70% of total barley SPLs undergoes alternative splicing which produced transcripts and proteins of different lengths. Further, protein-protein interaction network identified 35 potential interactors of barley SPLs that include mainly transcription factors and transporters. Only six SPL genes contained putative miR156 target site present in coding region except one gene where target site was found in 3′UTR region. Spatiotemporal expression pattern of the miR156- targeted SPLs showed tissue specific differential expression as compared to non-targeted SPLs. Transcript level of certain SPLs was found to be lower in the juvenile phase and higher in the reproductive phase of barley. Antagonistic expression pattern of miR156 and miR172 during the juvenile and the reproductive phases signifies their role in barley growth phase transition. Rajiv K. Tripathi [email protected]

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240-DV: Insights into the role of jasmonate signaling in the regulation of wheat seed dormancy

Tran Nguyen NGUYEN1, Belay T. AYELE1

1. Department of Plant Science, University of Manitoba, Winnipeg, Manitoba, Canada

Seed dormancy and germination are regulated by several plant hormones including abscisic acid (ABA), gibberellins (GA), ethylene and jasmonates (JA) and their interaction as well. To gain insights into the role of JA signaling in the regulation of dormancy and germination of wheat seeds, the transcript levels of JA signaling genes including JASMONATE ZIM-DOMAIN PROTEIN2 (JAZ2) and MYC TRANSCRIPTION FACTOR2 (MYC2) were measured in dormant and non-dormant wheat seeds at whole and different tissue levels. At whole seed level, similar expression level was observed between dormant and non-dormant seeds for JAZ2 and MYC2 before the start of imbibition. During imbibition, while MYC2 exhibited no difference, JAZ2 had a higher level of expression in non-dormant than dormant seeds. The difference in JAZ2 expression between dormant and non-dormant embryos was similar to that observed at the whole seed level, however, this expression difference was not apparent in the endosperm. With respect to the MYC2 gene, embryos from dormant seeds showed a considerably higher level of expression mainly during the later stage of imbibition than those derived from the non-dormant seeds. These results suggest embryo-specific role of JAZ2 and MYC2 in regulating dormancy maintenance and decay.

Tran Nguyen Nguyen [email protected]

241-DV: Analysis of interactions by the putative core stomatal proteins SPCH, MUTE and FAMA from Vitis M. Atikur RAHMAN1,2,$, Alison EDGE1,$, Ali EBADI1,3, Layla ALIBABAI1, Annette NASSUTH1 1. Department of Molecular and Cellular Biology, University of Guelph, Guelph, ON 2. Current Address: Agronomy Department, IFAS, University of Florida, Gainesville, FL 3. Department of Horticulture, University of Tehran, Karaj, 31578 Iran $. These authors contributed equally to this work The Arabidopsis bHLH proteins SPCH, MUTE and FAMA are key to the development of stomata. Their function relies on the formation of dimers with one of two other bHLH proteins called Arabidopsis ICE/SCRM1 and 2. We identified in Vitis one homolog each for SPCH, MUTE and FAMA, and four for ICE. The goal for this study was to determine which Vitis ICE proteins, if any, interact with which Vitis core stomatal protein. Dimer formation was investigated by BiFC, using agroinfiltrated N. benthamiana leaves transiently expressing various Vitis constructs. Optimized agroinfiltration conditions did not show any BiFC fluorescence for the negative control combination of ICE with CBF5, a nuclear localized, non-bHLH transcription factor. It was found that all Vitis core stomatal proteins form homodimers which translocate to the nucleus, except for MUTE-MUTE dimers which stay in the cytosol. Heterodimers were detected in the nuclei of cells expressing a combination of any of the stomatal and ICE proteins. However, for several combinations the percentage of CBF-RFP +ve nuclei which also were BiFC-positive was lower, suggesting that some dimers are less likely to form than others. Re-localization of MUTE-RFP from the cytosol to the nucleus if combined with any of its putative dimer partners supports the BiFC results. Current experiments investigate dimer formation by alternative core stomatal proteins, as encoded by detected alternative transcripts, and by FAMA mutated in its proposed dimerization domain. Annette Nassuth [email protected]

242-DV: Dormancy maintenance in wheat seeds is associated with repression of starch catabolism Menghan SUN, Yuji YAMASAKI, Feng GAO and Belay T. AYELE Department of Plant Science, University of Manitoba, Winnipeg, Manitoba, Canada Seed dormancy is an adaptive trait that blocks germination under apparently optimal conditions. Seeds can be released from the state of dormancy by various treatments including after-ripening. Loss of dormancy is associated

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with alterations in seed hormonal levels and sensitivity during imbibition, and this leads to changes in the expression of genes involved in many biological processes that regulate the germination process such as mobilization of storage reserves. Of the several plant hormones, gibberellins (GA) and abscisic acid (ABA) are the major players in controlling dormancy and germination, and therefore the associated biological processes. To better understand the molecular basis of starch degradation in wheat seeds with respect to dormancy maintenance and decay, and its regulation by GA and ABA, this study performed comparative expression analysis of starch degrading genes dormant and non-dormant (after-ripened) seeds. Dormancy loss due to after-ripening induced the expression of a number of starch degrading genes encoding amylases (AMY), α-glucosidase (AGL) and phosphorylase (PHO) genes during imbibition while dormancy maintenance was associated with the repression of these genes. The expression pattern of the starch degrading genes in both seed samples was closely associated with that of the GA biosynthetic genes. It appears from our result that ABA delays the germination of wheat seeds partly through repressing starch degrading genes.

Belay Ayele [email protected] 243-DV: TIR1 and AFB2 auxin co-receptors in developing pea fruit

Charitha P. A. JAYASINGHEGE, Jocelyn A. OZGA, Coutney D. NADEAU, Dennis M. REINECKE

Plant BioSystems Group, Department of Agricultural, Food, and Nutritional Science, University of Alberta, Edmonton, AB, Canada T6G 2P5 Pea fruit development requires the presence of seeds. Deseeded pericarps treated with naturally occurring pea auxin 4-chloro-indole-3-acetic acid (4-Cl-IAA) mimics the role of seeds and rescues fruit (pericarp) growth, and the other naturally occurring auxin indole-3-acetic acid (IAA) does not. To examine if the differential effect of these auxins on pea pericarp growth might be mediated in part through TIR1/AFB auxin co-receptors, we cloned the pea homologs of the TIR1 (PsTIR1a, PsTIR1b) and AFB2 genes and functionally characterized these gene in Arabidopsis. To determine whether the protein products of these putative pea auxin co-receptor genes were functional, these genes were expressed in Arabidopsis auxin receptor mutants Attir1-10 and/or Attir1-10afb2-3, under the regulation of the AtTIR1 promoter. Each pea gene restored the auxin response to mutant Arabidopsis seedling root growth. Gene expression analysis in pea pericarps showed that removal of seeds elevated pericarp PsTIR1b transcript abundance. Application of 4-Cl-IAA to deseeded pea pericarps reduced the transcript abundance of PsTIR1b, but IAA did not. Deseeding and auxin application minimally affected pericarp PsTIR1a and PsAFB2 transcript abundance. The auxin signaling pathway was functional in deseeded pericarps treated with either 4-Cl-IAA or IAA, as DR5-GUS gene expression was detected in deseeded pericarps (from DR5-GUS expressing pea plants) treated with either auxin. These data demonstrate that pericarp PsTIR1b gene expression is modulated by seeds and 4-Cl-IAA, but not IAA, and suggest that the auxin co-receptor pool may be important in bringing about the differential effects of the two auxins in developing pea fruits. Presenter: Dennis Reinecke [email protected] 244-DV: Stability and adaptability of peach cultivars in a subtropical climate Guilherme Locatelli1, Filipe Bittencourt Machado de Souza1, Rafael Pio1, Lee Kalcsits2, Rayane Barcelos Bisi1, Maraisa Hellen Tadeu1 1. Department of Plant Science, Federal University of Lavras, 37200-000 Lavras, Minas Gerais State, Brazil. 2. Washington State University Tree Fruit Research and Extension Center, Wenatchee, WA Prunus persica (peach) originated from a temperate climate and normally requires significant chilling to overcome dormancy. However, due to breeding, cultivars have been selected that can be grown in subtropical and tropical regions where there is low chilling unit accumulation during the dormant period. This study evaluated the adaptability and stability of the vegetative and reproductive phases of peach cultivars grown in a subtropical climate. An experiment was designed in randomized blocks with split plots, with 23 peach tree cultivars and four replications. To evaluate trait stability among cultivars, years and cultivars were considered to be treatments. The relative rates of budburst (RRB), flowering (RRF), shoot formation (SF) and fruit set (FS), average time to budburst (ATB) and flowering (ATF), duration of budburst (DB) and flowering (DF) and number of chilling hours (CH) with

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temperatures below 7.20 °C and 12 °C were measured during the experimental period. Fewer CH occurred during 2014, there were 279 hours below 12 ºC versus 436 hours below 12 ºC in 2015, and only five hours below 7.2ºC versus 37 hours below 7.2ºC in 2015. Based on the results, only two principal components sufficiently explain the evaluated variables, with proportions of 83% for SF and 76% for the RRB for the total variation. The cultivars ‘Bonão’, ‘Ouromel-4’, ‘Libra’ and ‘Maciel’ demonstrated greater adaptability during the vegetative phase, whereas ‘Ouromel-4’ and ‘Libra’ demonstrated greater adaptability and stability during the reproductive phase of peach trees in a subtropical climate. Keywords: Prunus persica, fruit set, shoot formation, chill requirement. Guilherme Locatelli [email protected]

245-DV: Two naturally auxins differentially modulate ethylene biosynthesis and response in the pea fruit. Charitha P.A. JAYASINGHEGE, Jocelyn A. OZGA, Kosala D. WADUTHANTHRI, Dennis M. REINECKE Plant BioSystems Group, Department of Agricultural, Food, and Nutritional Science, University of Alberta, Edmonton, AB, Canada T6G 2P5 Indole-3-acetic acid (IAA) and 4-chloroindole-3-acetic acid (4-Cl-IAA) are two naturally occurring auxins in pea. Only 4-Cl-IAA mimics the presence of seeds in the pea pericarp by stimulating GA biosynthesis and deseeded pericarp growth. Auxins also interact with ethylene. Seed and auxin regulation of pericarp ethylene biosynthesis and signaling was studied to explore why both auxins stimulate pericarp ethylene evolution similarly, but only 4-Cl-IAA inhibits ethylene response. In ethylene biosynthesis, S-adenosylmethionine is converted to 1-aminocyclopropane-1-carboxylic acid (ACC) by ACC synthase. ACC oxidase (ACO) converts ACC to ethylene. Regulation of the transcript abundance profiles of pericarp ACS (code for ACC synthases) and ACO (code for ACO oxidases) genes by 4-Cl-IAA was markedly different from that of IAA. 4-Cl-IAA dramatically increased PsACS1 transcript abundance up to 1800-fold (2 h after treatment), IAA induction was only 40-fold. Consistently, free ACC levels in 4-Cl-IAA-treated pericarps were about 20-fold higher than that in IAA-treated pericarps. PsACO genes were also modulated differently by the two auxins, but only minor changes in ACO enzyme activity were observed. Auxin-stimulated ethylene evolution was countered by 4-Cl-IAA-specific decrease in ethylene responsiveness potentially via modulation of pericarp ethylene receptor and signaling gene expression. This pattern did not occur in IAA-treated pericarps. Overall, the effect of 4-Cl-IAA and IAA on ethylene biosynthesis gene expression generally explains the ethylene evolution patterns, and their effects on GA biosynthesis and ethylene signaling gene expression explain the tissue response patterns in young pea ovaries. Presenter: Jocelyn Ozga [email protected]

246-DV: Proteogenomics analysis of seed dormancy identifies genotype- and phenotype-associated proteomic signatures of pre-harvest sprouting resistance in dormant and non-dormant hybrid genotypes of wheat Natalia V. BYKOVA1, Mark JORDAN1, Nataša RADOVANOVIC1, Michelle RAMPITSCH1, Junjie HU1,2 1. Morden Research and Development Centre, Agriculture and Agri-Food Canada, Morden, MB 2. Department of Biology, Memorial University of Newfoundland, St. John's, NL Control of pre-harvest sprouting (PHS) resistance in wheat is complex, genetic factors responsible for it are dispersed on almost every wheat chromosome and interactions occur between QTLs and/or among environments. Elucidation of molecular mechanisms controlling seed dormancy, after-ripening and environmental regulation reveal sources of genetic variation. In this study, we used proteogenomics approach to identify seed tissue-specific proteomic signatures of PHS resistance. Dormancy genotype- and phenotype-associated alterations in aleurone and embryo proteomes were analyzed using a hard white spring wheat (Triticum aestivum L.) hybrid doubled haploid lines with marginal dormancy phenotypes, iTRAQ-based proteomics integrated with customized EST database, high-resolution nanoLC-MS/MS and Scaffold Q+ quantitative analysis, Mercator annotation pipeline, MapMan pathway mapping, and association with QTL regions. Altogether 6810 proteins in 4116 clusters were identified with

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high confidence, of which 62 and 115 unique proteins showed significant differential expression in dormant phenotypes, and 368 and 1041 unique proteins were dormancy genotype-specific in embryo and aleurone, respectively. In dormant embryos, significant phenotype-specific changes were found for proteins involved in redox controlling system, signaling associated with flowering, development and growth repression, hormone signaling, cell cycle control and epigenetic regulation of gene expression, cell wall metabolism, vesicle transport, and ubiquitin 26S proteasome pathway. Phenotype-specific changes in aleurone involved proteins responsible for defenses against pathogens, energy metabolism, protein turnover, RNA processing, nucleotide synthesis, vitamin metabolism, G-proteins, ABA and ethylene metabolism. Corresponding genes on chromosome arms where QTL for PHS tolerance had been previously identified were further analyzed to compare their location in the QTL region. Natalia Bykova [email protected]

247-DV: Plant hormone and MAPK signalling pathways in wheat

Nora A. FOROUD1, Ravinder K. GOYAL1, André LAROCHE1, Brian E. ELLIS2, Dan TULPAN4, Ziying LIU4, Youlian PAN4

1. Agriculture and Agri-Food Canada, Lethbridge Research and Development Centre, Lethbridge, AB, Canada 2. Michael Smith Laboratories, University of British Columbia, Vancouver, BC, Canada 3. National Research Council of Canada, Ottawa, ON, Canada

Mitogen-activated protein kinases (MAPKs) and plant hormones are important signalling molecules regulating plant development and defence responses. Cross-talk between these two signalling pathways has been reported in plants. Despite their importance in biological processes the MAPK signalling cascades are poorly characterized in important cereal crops, such as wheat. We recently surveyed the genomic DNA database in Ensembl Plant and identified the MAPK members of the MAP kinase (MPK) and MAPK kinase (MKK) subfamilies in wheat and related species. Seventeen MPKs and 12 MKKs, along with multiple genomic copies, were identified in hexaploid wheat. Orthologous naming was assigned based on the accepted MAPK nomenclature to facilitate comparisons with other plant species. Compared with MAPKs, hormone signalling pathways have received considerably more attention in wheat, and increasing evidence indicates that these pathways differ in cereals compared with dicots and, to a lesser extent, other monocots. In an RNA-sequencing experiment, differential gene expression was analyzed in wheat spikes treated with salicylic acid, methyl-jasmonate, or ethephon. The wheat MAPK sequences were mapped onto this data-set to observe how hormone treatments affected expression of MAPK genes. The methyl-jasmonate treatment was most effective in eliciting expression of numerous MAPKs in wheat spikes and no induced changes were observed in response to salicylic acid. Experiments are currently underway to determine whether the MPK enzymes are differentially activated in response to these three hormones.

Nora Foroud [email protected]

248-DV: DEMETER as a possible link between DNA methylation status and homologous recombination in Arabidopsis

Tawhidur RAHMAN1, Kevin Rozwadowski2

1Department of Plant Sciences, College of Agriculture and Bioresources, University of Saskatchewan, Saskatoon, SK, Canada S7N 5A8 2Agriculture and Agri-Food Canada, Saskatoon, SK, Canada S7N OX2 DEMETER (DME) is a bifunctional DNA glycosylase/apyrimidic lyase that was originally identified as an epigenetic regulator of seed development in Arabidopsis whereby it activates a subset of regulatory genes by catalyzing the removal of repressive 5-methylcytosines at CpG sites in their promoters. While investigating DNA recombination processes in plants, we identified DME as a physical interactor of RAD51, a highly conserved mediator of homologous recombination in eukaryotes. This physical interaction was delineated to a unique intrinsically disordered region of DME and an N-terminal region of RAD51. DME and RAD51 displayed likewise

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patterns of induced expression in response to exposure to DNA damaging agents, including gamma radiation, bleomycin and camptothecin. Since dme seeds are non-recoverable due to embryonic lethality caused by homozygosity of the null mutation, we developed a novel conditional complementation strategy to establish lines that are DME-proficient during embryo development but subsequently functionally dme-null, as shown by lack of DME transcript and hypermethylation of MEDEA, an epigenetically regulated target of DME. Conditionally complemented dme and rad51-null lines were hypersensitive to DNA damage mediated by camptothecin and bleomycin, providing genetic evidence for DME having a role in DNA repair processes and functioning in the same pathway as RAD51. The coincident action of DME and RAD51 in DNA repair was further supported using respective promoter fusions to GFP, which revealed coordinate induction kinetics for both genes in root cells in response to gamma irradiation and camptothecin. Collectively, our results implicate DME as a new component of DNA repair responses in plants and broaden its role beyond epigenetic regulation of imprinted genes, as well as provide novel insight to modulation of DNA methylation status as being a possible factor in homologous recombination-mediated DNA repair. Tawhidur Rahman; email – [email protected] / [email protected]

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Education Posters (249-‐ED to 250-‐ED) Thursday July 6, FSC Atrium 11:20am -12:30pm 249-ED: Example Answers Support Problem Solving in Second- and Third-Year Biology Courses

Miranda J. Meents1, Robin E. Young1, Sunita Chowrira1, A. Lacey Samuels1 1 Department of Botany, University of British Columbia, Vancouver, Canada One of the most important skills that undergraduate students in the sciences learn are problem solving skills. One strategy instructors frequently use in supporting students learning these skills is asking students to solve practice problems. For biology classes, this often involves using analysis and interpretation of data to generate a clear and concise hypothesis, and then defend it using evidence and logical reasoning. However, mastering each of these skills and integrating them to successfully solve a problem can be extremely difficult for novices. In many disciplines students have been successfully supported in their problem-solving efforts using example answers, which help guide students through the problem-solving process. We tested the effect of providing example answers on student learning in two undergraduate cell biology courses using a combination of student surveys, feedback, and grades on problem solving tasks. The results show that example answers can improve student problem solving in some cases. Students also consistently felt better supported in their efforts when given example answers. Importantly, the benefits of example answers increased for ‘average’ students, compared to exceptional students. Together these results indicate that example answers can be a valuable component of teaching biological problem solving to undergraduate students, especially if other forms of support are limited.

Presenting Author: Miranda Meents [email protected] 250-ED: Enrichment of plant physiology lab education by course-based undergraduate research experiences (CURE) Santokh SINGH1* 1. Department of Botany, University of British Columbia, Vancouver, BC, Canada V6T 1Z4 In an effort to enhance the educational enrichment experience in two upper-level plant physiology courses, course-based undergraduate research experiences (CURE) and Three Minute Thesis (3MT)-based oral presentations were introduced. Undergraduate research students work in small groups and develop a research proposal. They design and conduct inquiry-driven, open-ended research projects in the areas of plant physiology, plant development and biochemistry. Students conduct a series of relevant research experiments, and analyze their experimental data. Each student communicates his/her research findings in the form of written research reports according to the publishing guidelines of the Plant Physiology journal. In addition, I have introduced the “Three Minute Thesis (3MT)” model of short research presentations as an education tool and assessment method in these courses. Finally, each student gives a short research presentation similar to the “Three-Minute Thesis (3MT) format. The 3MT presentations improved the scientific communication and research skills of students. The significance and effectiveness of course-based undergraduate research experiences (CURE) and Three-Minute Thesis (3MT) will be discussed. Santokh Singh [email protected]

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Specialized Metabolism Posters (251-‐SM to 260-‐SM) Thursday July 6, FSC Atrium 11:20am -12:30pm 251-SM: The waxes covering sword fern (Polystichum munitum) fronds: a complex mixture of alkyl esters and estolides reminiscent of gymnosperms and angiosperms

Alberto RUIZ ORDUNA1*, Yanjun GUO2, Lucas BUSTA1, Reinhard JETTER1,2 1. Department of Chemistry, University of British Columbia, Vancouver, BC 2. Department of Botany, University of British Columbia, Vancouver, BC The early steps of land colonization required plants to evolve a hydrophobic coating for protection against environmental stress imposed by the atmosphere, most importantly to prevent excessive loss of water. Ferns are among the earliest vascular plant species, and comparisons between their cuticle structure and composition with those of angiosperms may thus give information about the ancestral land plant cuticle. Here, the cuticular wax covering sword fern (Polystichum munitum) fronds was analyzed to identify novel compounds and to quantify all components within the complex mixture. Large amounts of alkyl esters were detected (ca. 80% of the total wax coverage), spanning a wide range of C36 to C60 homologs. Based on mass spectrometric fragmentation patterns and comparisons with synthetic standards, ester metamers were found composed of both long-chain (C16/C18) and very-long-chain (C20+) primary alcohols and fatty acids, similar to angiosperm wax esters. A second ester fraction was identified as trimers linking a C14/C16 α,ω-diol with two C12 - C16 fatty acids. Interestingly, similar estolide structures had been reported sporadically before, mostly for gymnosperms. The vast variety of wax esters in sword fern wax suggests the presence of several wax ester synthase (WS) enzymes with distinct chain length specificities. It will be very interesting to further characterize those enzymes, to expand our current knowledge on WSs and their evolution.

Alberto Ruiz Orduna: [email protected] 252-SM: The Arabidopsis thaliana Rhizobiale-like phosphatase 2 is a novel D-group MAPK tyrosine-specific PPP-family protein phosphatase.

Anne-Marie LABANDERA1*, Glen UHRIG2, Ryan TOTH1, Keaton COLVILLE1, Kenneth K.S. NG1, Greg G.B. MOORHEAD1 1. Department of Biological Sciences, University of Calgary, Calgary, AB, Canada 2. Plant Biotechnology, Department of Biology, Swiss Federal Institute of Technology (ETH), Zurich, Switzerland. The study of tyrosine phosphorylation in plants has been largely neglected due to the lack of classic tyrosine kinases and underrepresentation of tyrosine phosphatases compared to humans. However, advanced phosphoproteomics studies have revealed that the abundance of phospho-tyrosine residues in plants parallels humans. This strongly suggests that in plants tyrosine phosphorylation is as important as in humans, yet we know nothing about the players responsible of these events. The Arabidopsis thaliana Rhizobiale-like phosphatase 2 (AtRLPH2) is a novel protein phosphatase not found in mammals which, according to bioinformatics analysis, clusters with the serine/threonine specific phospho-protein phosphatase (PPP) group. In the present work, we demonstrate that AtRLPH2 surprisingly behaves like a tyrosine phosphatase. AtRLPH2 is not affected by classic PPP inhibitors, but in turn is inhibited by the specific tyrosine phosphatase inhibitor, sodium orthovanadate. Moreover, AtRLPH2 dephosphorylates phospho-tyrosine substrates and has essentially no activity towards phospho-serine/threonine residues. These evidences show for the first time that a member of the plant PPP family of phosphatases has the capability to dedicate its activity solely toward phospho-tyrosine. Furthermore, in order to identify AtRLPH2 substrates, a phosphoproteomics study was performed by comparing phospho-tyrosine peptides from wild type (WT) and atrlph2 knockout plant lines. AtRLPH2 was found and confirmed to be a D-group specific Mitogen Activated Protein Kinase (MAPK) phosphatase. Moreover, we elucidated AtRLPH2 crystal structure providing detailed mechanistic insight into its mode of action and substrate specificity.

Anne Marie Labandera [email protected]

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253-SM: Detoxification of Pine Terpenoids by the Mountain Pine Beetle Christine CHIU1*, Christopher I. KEELING1,3, Jörg Bohlmann 1 1. University of British Columbia, Michael Smith Laboratories, 2185 E Mall, Vancouver, BC V6T 1Z4 2. Botany Department, University of British Columbia, 6270 University Blvd, Vancouver, B.C., V6T 1Z4 3. Present address: Department of Biological Sciences, Simon Fraser University, 8888 University Drive, Burnaby, B.C., Canada, V5A 1S6

Since the late 1990s, over 25 million hectares of lodgepole pine (Pinus contorta) forests have been affected by a continuous outbreak of the mountain pine beetle (Dendroctonus ponderosae; MPB), a bark beetle pest native to North America. This outbreak has seen the extension of the MPB range across the previous barrier of the Rocky Mountains, into a novel habitat of jack pine (Pinus banksiana), which continues through Canada’s boreal forest. Pines produce mono-, sesqui- and diterpenoids as components of the oleoresin, an important chemical and physical defense against insect and pathogen attack. MPB is able to tolerate and detoxify high concentrations of the terpenoids and also uses a pine monoterpene as a precursor to its aggregation pheromone trans-verbenol. Through a functional genomics approach, we have uncovered several MPB cytochromes P450s that detoxify pine terpenoids. We have quantified the toxic effects of monoterpenes on MPB and identify a novel mechanism by which MPB both detoxifies and sequesters monoterpenes. We show the role that MPB P450s have in transforming pine monoterpenes through enzyme activity assays with recombinant P450s, transcript levels (qPCR), and gene silencing (RNAi) of P450s in MPB. Christine Chiu [email protected]

254-SM: Creating a synthetic microbial platform for diterpenoids production Joseph C. UTOMO1,*, Moonhyuk KWON1, Sihyun BAEK1, Fabio CHAVES1, Vincent J.J. MARTIN2, Dae-Kyun RO1* 1. Department of Biological Sciences, University of Calgary, Calgary, AB, Canada 2. Department of Biology, Concordia University, Montreal, QB, Canada Diterpenoid is a subclass of terpenoid superfamily, all of which are synthesized from a simple precursor isopentenyl diphosphate (IPP). Diterpenoids possess 20-carbon backbones, and their direct precursor is C20 geranylgeranyl diphosphate (GGPP). Many pharmaceuticals and nutraceuticals belong to diterpenoids, and a number of the biosynthetic genes for medically important diterpenoids, such as taxol, have been identified in recent years. However, practical tools to aid the bulk production of diterpenoids are still lacking, hindering the research on diterpenoids metabolisms. To achieve a versatile microbial platform to produce various diterpenoids, we chose yeast (Saccharomyces cerevisiae) for metabolic modifications with an aim to produce diterpenoids taxol precursor, taxadiene. Taxadiene synthase (TmTDS) cDNA was isolated from Taxus media (Anglojap yew), and its expression in yeast showed a nano-gram scale of taxadiene in L-culture. Naturally yeast does not have any GGPP-derived metabolites, and its GGPP synthase is primarily used to produce a minute quantity of GGPP for protein prenylations. Therefore, to increase the level of GGPP in yeast, we built and used an IPP-producing plasmid encoding 4 genes in the mevalonate pathway (called IPP-plasmid) and also overexpressed yeast GGPP synthase. Incorporating IPP-plasmid and yeast GGPP synthase overexpression in the base strain (i.e., overexpressing TmTDS alone) resulted in 212-fold increase of taxadiene. We are currently testing GGPP synthases from different organisms (bacterium, algae, and plant) and altering yeast genome by a CRIPSR genome-editing tool to further improve taxadiene production in yeast. Presenter: Joseph C. Utomo [email protected]

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255-SM: Identification of transcription factors that regulate terpene biosynthesis in lavenders Lukman S. SARKER1*, Soheil MAHMOUD 1. Department of Biology, University of British Columbia (Okanagan Campus), BC, Canada

Lavenders are aromatic shrubs known for their fragrant essential oils (EO), which are dominated by mono- and sesquiterpenes. The biosynthesis of lavender EO constituents begins with the synthesis of universal terpene building blocks isopentenyl diphosphate (IPP) and its allylic isomer dimethylallyl diphosphate (DMAPP). The enzymes called terpene synthases (TPSs) are involved in the conversion of these precursors into EO constituents. Although many lavender terpene synthases have been cloned and characterized, a limited amount of information is available regarding the transcriptional regulation of TPS genes in lavender. This study aims to shed light on the regulation of terpene biosynthesis by identification of transcription factors (TFs) that control the expression of TPSs in lavenders. Using the Illumina platform, approximately 30 million high quality reads were obtained from flower and leaf RNA of three economically important lavender species. The de novo assembly resulted in a transcriptome with more than 101,000 unigenes, 75% of which were annotated using public databases. More than 2000 transcription factor (TF) candidates were identified from the transcriptome database, and 35 candidates were selected- based on their quantitative expression- for further analysis. This study will enhance our understanding of regulation of terpene metabolism in lavenders.

Lukman Sarker – [email protected]

256-SM: Probing Cannabis sativa glandular trichome cellular ultrastructure during resin production Samuel J. LIVINGSTON*1, Teagen D. QUILICHINI2, Eva Y. CHOU1,3 Jon E. PAGE1,3, A. Lacey SAMUELS1 1. Department of Botany, University of British Columbia, Vancouver, BC 2. National Resource Council Canada, Saskatoon, SK 3. Anandia Laboratories, Vancouver, BC Cannabis sativa L. has glandular trichomes, which protrude from the epidermis of floral tissues of the female plant and produce a diverse array of lipophilic metabolites. Despite the social and economic importance of cannabis, how the organelles of the highly-metabolic disk cells of the glandular trichomes produce the abundant lipid metabolites is unknown. The goal of this study was to reveal the features of disk cell ultrastructure related to their specialized metabolism. Conventional chemical fixation (CF) and high-pressure freezing followed by cryofixation (HPF) were used to preserve cellular ultrastructure for transmission electron microscopy (TEM). Consistent features observed using the two techniques include highly polarized cytoplasmic contents, where plastids and mitochondria are enriched at the apical domain of disk cells, with vacuoles and nuclei predominantly at the basal domains. In CF-glands, the cells contained amorphous oil body-like inclusions, notably on the surface of plastids and mitochondria, as well as between the plasma membrane and cell wall. However, inclusions were not observed in glands preserved with HPF, suggesting that the inclusions may be artifacts of lipid remobilization during fixation. In contrast, HPF samples contained abundant smooth endoplasmic reticulum, membrane contact sites, and plastids with paracrystalline cores and complex internal membranes. Vacuolar morphology in CF-prepared samples was irregular in shape and distinguished by a highly osmiophilic margin, whereas HPF-prepared samples possess a predominantly spherical vacuole with a homogenous electron-dense composition. This study reveals the importance of selecting an appropriate method of cellular preservation for investigating the ultrastructure of tissues with abundant lipid metabolism. Samuel J. Livingston [email protected]

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257-SM: Identification of loci for volatiles accumulation in tomato using an introgression line population derived from Solanum lycopersicoides Marie-Ève MONTMINY1, Jonathan TREMBLAY1, Louis-Félix NADEAU1, Blandine BULOT1, Charles GOULET1 1. Centre de recherche et d’innovation sur les végétaux, Département de phytologie, Université Laval, Québec, QC Introgression lines have been used to discover quantitative trait loci (QTL) in tomato for numerous traits. These lines who carry small genome segments of wild species have notably allowed the identification of genes controlling volatiles production in tomato (Solanum lycopersicum). In this study, we use a population of S. lycopersicoides introgression lines to identify new QTL associated with volatiles accumulation in tomato fruits. S. lycopersicoides is the furthest species that can be crossed with S. lycopersicum with a divergence event estimated at 9 million years ago. This great genetic distance provides a chance to discover QTL that were not observed in introgression lines derived from closest species. Sixty lines covering all chromosomes were grown in the field for several years and volatiles were collected to identify the QTL. In total, more than thirty genome regions were linked with volatiles accumulation, notably many volatiles derived from branched-chain amino acids and fatty acids. The QTL identified will be used to further identify genes that modulate volatiles production and to better understand the genetic behind tomato flavour. Charles Goulet, [email protected]

258-SM: A WRKY transcription factor from Withania somnifera regulates triterpenoid withanolides accumulation and biotic stress tolerance through modulation of phytosterol and defense pathways

Anup K. SINGH1, Sarma R. KUMAR1, Varun DWIVEDI1, Avanish RAI1, Shaifali PAL2, Ajit K. SHASANY2 and Dinesh A. NAGEGOWDA1

1. Molecular Plant Biology and Biotechnology Lab, CSIR-Central Institute of Medicinal and Aromatic Plants Research Centre, Bengaluru - 560065, India

2. Biotechnology Division, CSIR-Central Institute of Medicinal and Aromatic Plants, Lucknow - 226015, India

Withania somnifera (L.) Dunal produces pharmacologically important triterpenoid steroidal-lactones known as withanolides, whose accumulation is induced by certain phytohormones indicating the involvement of transcriptional control network. To investigate the transcriptional regulation of withanolides biosynthesis, we have characterized the biological role of W. somnifera WRKY1 (WsWRKY1), a group-III WRKY transcription factor that is induced in response to methyl jasmonate and salicin, predominantly expressed in leaves, and localizes to the nucleus. Downregulation of WsWRKY1 by virus-induced gene silencing (VIGS) in W. somnifera resulted in stunted plant growth, reduced transcripts of phytosterol pathway genes with corresponding reduction in phytosterols and withanolides. In contrast, transient overexpression of WsWRKY1 in W. somnifera leaves up-regulated sterol pathway genes and enhanced the accumulation of phytosterols and withanolides. Moreover, ectopic expression of WsWRKY1 in tobacco led to up-regulation of sterol pathway genes with significant increase in phytosterols. Chromatin immunoprecipitation (ChIP) assay showed that WsWRKY1 binds to W-box sequences in promoters of W. somnifera genes encoding squalene synthase and squalene epoxidase, indicating its direct regulation of triterpenoid pathway. Furthermore, while WsWRKY1 silencing in W. somnifera compromised the tolerance to bacterial growth, fungal infection, and insect feeding, its overexpression in tobacco led to improved biotic stress tolerance. Together these findings demonstrated that WsWRKY1 has a positive regulatory role on phytosterol and withanolides biosynthesis, and defense against biotic stress, highlighting its importance as metabolic engineering tool for simultaneous improvement of triterpenoid biosynthesis and plant defense. [email protected]

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259-SM: Identification and characterization of an E3 ubiquitin ligase that negatively regulates cuticular wax biosynthesis in Arabidopsis Shuang LIU1*, Meixuezi TONG1,2, Xin LI1,2, Ljerka KUNST1 1. Department of Botany, University of British Columbia, Vancouver, CA 2. Michael Smith Laboratories, University of British Columbia, Vancouver, CA Cuticular wax is a key component of the plant cuticle and its accumulation is known to be tightly regulated transcriptionally and post-transcriptionally in response to developmental and environmental cues. Recently, we conducted an E3 ubiquitin ligase reverse genetic screen and identified a RING-type E3 ligase, designated U37, which negatively regulates wax biosynthesis in Arabidopsis thaliana. Plants overexpressing U37 display glossy stems and siliques, as well as reduced fertility and fusions between aerial organs. Total stem wax load in U37 overexpression lines was reduced by >70%, with proportional decreases in alkanes, secondary alcohols, and ketones, and increases in aldehydes, acids, and primary alcohols, suggesting that the wax biosynthetic enzymes and/or positive regulators in the alkane-forming pathway may be the potential target(s) of U37. Overexpression of a dominant-negative version of U37 protein had no effect on the wax load. Transient co-expression of U37 and its candidate targets in Nicotiana benthamiana leaves revealed that CER1 was degraded via the 26S proteasome pathway. The U37-dependent turnover of CER1 has been confirmed in stable transgenic lines of Arabidopsis. Similar to CER1, the degradation of CER3 in the presence of U37 has also been detected in the N. benthamiana transient expression system. The discovery of U37 demonstrates that post-translational regulation is also involved in cuticular wax deposition. Further studies of U37 will shed new light on the regulation of wax biosynthesis in Arabidopsis. Shuang Liu [email protected]

260-SM: Arabidopsis ECERIFERUM2-LIKEs are mediators of condensing enzyme activity Tegan M. HASLAM, Wesley K. GERELLE, Sean W. GRAHAM, Ljerka KUNST Department of Botany, University of British Columbia, Vancouver, BC

Cuticular waxes seal the primary aerial surfaces of land plants, providing a robust water barrier that protects against desiccation. Cuticular waxes are predominantly acyl-lipids ranging from 24-36 carbons in length, and are derived from very-long-chain fatty acids (VLCFAs). The core elongase complex that synthesizes VLCFA wax precursors consists of four enzymes that reside on the endoplasmic reticulum. Condensing enzymes catalyze the committed reaction of elongation, and determine the chain length of fatty acid substrate accepted by the complex. While necessary for the elongation of all VLCFAs, identified plant condensing enzymes cannot efficiently produce VLCFAs longer than 28 carbons. This falls short of the requirements of wax biosynthesis. The eceriferum2 (cer2) mutant of Arabidopsis thaliana has a severe wax-deficient phenotype and specifically lacks waxes longer than 28 carbons, but the CER2 protein has no homology to condensing enzymes. Heterologous expression in yeast has demonstrated that CER2 can modify the chain-length specificity of a condensing enzyme, CER6, from 28 to 30 carbons. There are five CER2-LIKE genes in Arabidopsis, each of which has a different effect on condensing enzyme activity. Structural models, domain swaps, site-directed mutagenesis, and protein-protein interaction studies were used to elucidate the role of CER2-LIKEs in elongation. Several mechanisms are proposed to explain the biological function of CER2-LIKE proteins.

Tegan M. Haslam [email protected]

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Plants from Sea to Sky Delegates

Last Name First Name Affilliation Abbasi Pervaiz Agriculture and Agri-Food Canada 224-BI Abdulmajeed Awatif Dalhousie University AB1.6 Adal Ayelign

Mengesha UBC Okanagan SM1.4

Adhikary Dinesh University of British Columbia, Okanagan

SM2.2

Adihetty Dilini University of Alberta 238-DV Alharthy Azizah University of Manitoba 101-AB Ao Kevin University of British Columbia BS.4 Aslam Sher University of Haripur Awwad Fatima Centre Sève-Université de Sherbrooke CB2.4 Ayele Belay University of Manitoba Baek Sihyun University of Calgary Bahrani Hirbod U of S, Department of Plant Science AB2.6 Barcelos Bisi Rayane Washington State University 200-AP Barghahn Sina Georg-August-University Goettingen 137-BS Beatty Perrin University of Alberta NM.2 Beaudoin Guillaume University of Florida BC.5 Behar Hila University of British Columbia Belmonte Mark University of Manitoba BI.2 Benic Elena University of Saskatchewan 116-AB Benitez Francisco University of British Columbia 142-CB Besik Ariana University of Toronto Bilek Michael University of British Columbia 205-AP Bird David Mount Royal University CB2.3 Bisgrove Sherryl Simon Fraser University Blutrich Ron University of British Columbia 147-CB Bohlmann Joerg University of British Columbia Booth Judith University of British Columbia SM1.6 Bors Robert University of Saskatchewan 207-AP Bracken Matthew University of California, Irvine Plen2.2 Braeutigam Katharina University of Toronto Mississauga SR.5 Braybrook Siobhan University of Cambridge Plen1.2 Burlakoti Rishi Agriculture and Agri-Food Canada AP.6

123

Buschhaus Christopher Crandall University EP.1 Bykova Natalia Agriculture and Agri-Food Canada 246-DV Caldo Kristian Mark University of Alberta BC.4 Carella Philip Sainsbury Laboratory, University of

Cambridge BI.4

Castellarin Simone UBC Wine Research Centre AB2.1 Celedon Jose M University of British Columbia SM1.2 Chakraborty Sonhita University of Toronto BS.3 Charles Marie Thérèse Agriculture and Agri-Food Canada AP.7,

120-AB Chen Lynn University of British Columbia 234-CW Chen Melissa University of British Columbia BI.6 Chera Aman University of British Columbia Chiu Christine Michael Smith Laboratories 253-SM Clayton Emily UWO Clermont Isabelle Université Laval 202-AP Clinton Mary Simon Fraser University 107-AB Cloutier Sylvie Agriculture and Agri-Food Canada Collyer Danielle Simon Fraser University 218-BT Congreves Kate University of Saskatchewan NM.1 Constabel C. Peter University of Victoria SM1.1 Cronk Quentin University of British Columbia Da Ros Letitia University of British Columbia 106-AB Dastmalchi Mehran University of Calgary SM2.1 de Albuquerque Igor University of Saskatchewan 121-BS Dean Gillian University of British Columbia Debnath Samir Agriculture & Agri-Food Canada 208-AP Dejeto Leni University of the Philippines - Visayas

Tacloban College

Demissie Zerihun National Research Council of Canada 141-BS Desjardins Yves Laval University Plen2.3 Desta Demissew Sertse UOttawa/AAFC Dhillon Amneet University of British Columbia 115-AB,

201-AP Dias Kayla University of Toronto 105-AB Dimopoulos Nicholas University of British Columbia AB1.4 Ding Yuli University of British Columbia 128-BS Dominguez Sergio Western University 100-AB Donnelly Caitlin University of British Columbia Duarte Andre G The University of Western Ontario AB2.7

124

Edwards Diane ABI Environmental Services Ltd Ehlting Juergen University of Victoria CDS.2 Elferjani Raed Agriculture and Agri-Food Canada EP.4 Ensminger Ingo University of Toronto, UTM Biology TI.1 Feng Hui University of Saskatchewan Feyissa Biruk Ayenew University of Western Ontario AB2.4 Ficzycz Natasha Simon Fraser University 109-AB Fofana Bourlaye Agriculture and Agri-Food Canada AP.4,

130-BS Foroud Nora AAFC-Lethbridge 247-DV Frank Joshua Western University 220-BI Fujita Miki University of British Columbia 150-CB Geitmann Anja McGill University CB2.6 Ghahremani Mina Queen's University NM.4 Gonzales-Vigil Eliana University of British Columbia BC.3 Goring Daphne University of Toronto Goulet Charles Université Laval 257-SM Gourlay Gerry University of Victoria AB1.5 Gravel Valerie McGill University BI.1 Green Beverley University of B.C. 154-NM Groover Andrew US Forest Service & UC Davis CDS.1 Gutierrez Rodrigo University of British Columbia 204-AP Guy Rob University of British Columbia Guzha Athanas Georg-August University 129-BS Halat Laryssa University of British Columbia 145-CB Hamilton Kaila University of Saskatchewan 119-AB Haney Cara University of British Columbia Plen1.3 Haslam Tegan University of British Columbia 260-SM Hawkins Barbara University of Victoria NM.5 Hawley Dave University of Guelph TI.4 Hodgins Connor University of Calgary SM1.5 Hoffmann Natalie University of Toronto 123-BS Holguin Jorge University of British Columbia 143-CB Holloway David UBC, Math Dept Hossain MD Musharaf University of Saskatchewan BS.5 Hossain Sarzana University of British Columbia 222-BI Hu Hefeng (Jasmine) Carleton University EP.2 Hu Yi University of British Columbia Hu-Skrzenta Junjie Grace Memorial University of Newfoundland DV.5

125

Huang Weijie University of British Columbia 126-BS Huang Xinyi University of British Columbia 111-AB Hughes Scott University of Western Ontario 125-BS Igamberdiev Abir Memorial University of Newfoundland BC.2,

113-AB Indriolo Emily New Mexico State University 159-SR Irani Solmaz University of Saskatchewan 138-BS Ische Reilly University of British Columbia Jamsed Muhammad University of Calgary 160-SR Jetter Reinhard University of British Columbia BC.6 Jo Seohyun University of British Columbia SM2.3 Jones Melanie UBC Okanagan BI.3 Joshi Jaya Western University SS.1 Kalcsits Lee Washington State University TI.6 Kambhampati Shrikaar Western University 152-NM Kapoor Karuna University of Manitoba EP.3 Karia Purva University of Toronto 124-BS Kariyawasam Thamali University of British Columbia SR.4 Kaur Surinder SGTB Khalsa College, University of

Delhi 139-BS

Keerthisinghe Sandra University of British Columbia 151-CB Khan Deirdre University of Manitoba SS.2 Khan Madiha University of Toronto TI.5 Kim Hae Ryoung Simon Fraser University CB1.2 Klee Harry University of Florida, Gainesville Plen3.1 Kovalenko Yevgen University of British Columbia 210-AP Kshatriya Kristina University of British Columbia SS.4 Kumar Abhinandan University of Calgary 155-SR Kwon Moonhyuk University of Calgary Labandera Anne Marie University of Calgary 252-SM Lajeunesse Julie Agriculture and Agri-Food Canada 203-AP Lakusta Adam University of Calgary SM2.5 Lan Xingguo University of Calgary Le Yen University of Saskatchewan CB1.1 Lee EunKyoung UBC Botany 149-CB Lee Hyun Kyung University of Toronto SR.2 Lee Jae-Hyeok University of British Columbia DV.1 Lee Jin Suk Concordia University Lee Joon Seon University of British Columbia AB2.3

126

Lee Shinwoo Gyeongnam National University of Science & Technology

209-AP

Lee Yi-Chen University of British Columbia 148-CB LI KEKE University of Saskatchewan Li Meng University of British Columbia DV.2 Li Xin University of British Columbia Li Yuhua Northeast Forestry University Li Zhuowei

(Charles) University of British Columbia

Lin Jordan University of British Columbia 223-BI Liu Yanan University of British Columbia Liu Shuang University of British Columbia 259-SM Liu Yuanyuan University of British Columbia Liu Zhexian University of British Columbia 225-BI Livingston Samuel University of British Columbia 256-SM Locatelli Guilherme Federal University of Lavras, Brazil 244-DV Luedke Daniel Georg-August-University Goettingen 131-BS Luo Yanjie Macfie Sheila University of Western Ontario MacKay Heather University of British Columbia 212-BC Mahmoud Soheil University of British Columbia Mahon Elizabeth University of British Columbia 122-BS Maltais Anne-Marie Université Laval 215-BT Mansfield Shawn University of British Columbia Marquez Mellidez Carmen University of Saskatchewan 237-DV Martone Patrick University of British Columbia CB2.1 Matthews Craig University of Western Ontario 104-AB Mattsson Jim Simon Fraser University DV.4 McCotter Sean University of British Columbia McElrone Andrew University of California, Davis TI.2 McGee Robert University of British Columbia CB2.2 McGregor Nicholas University of British Columbia 235-CW McIsaac Matthew Thompson Rivers University 158-SR Meents Miranda University of British Columbia CB1.4,

249-ED Megha Swati University of Alberta 110-AB Melnyk Ryan University of British Columbia BI.5 Merchant Sabeeha University of California, Los Angeles Plen2.1 Meyerowitz Elliot Caltech Plen1.1 Micallef Barry University of Guelph

127

Milla-Moreno Estefania University of British Columbia Miller Shea Agriculture & Agri-Food Canada 136-BS Moffett Peter Université de Sherbrooke 135-BS Momayyezi Mina University of British Columbia NM.6 Monaghan Jacqueline Queen's University 133-BS Moorhead Greg University of Calgary BC.1 Mottair Yaseen University of British Columbia 232-CW Munz Jacob University of British Columbia 152-NM Murch Susan University of British Columbia AP.2 Murphy Emily University of British Columbia 217-BT Nagegowda Dinesh University of British Columbia 258-SM Nair M.P.M. LLT Plants Inc. AP.3 Nandi Shyamal G.B. Pant National Institute of

Himalayan Environment & Sustainable Development

SM2.4

Nassuth Annette University of Guelph AB1.2, 241-DV

Nelles Hayley University of Toronto 156-SR Nguyen Tran Nguyen University of Manitoba 240-DV Nguyen Trinh University of Manitoba SS.3 Nitta Yukino University of British Columbia Ouellet Thérèse Agriculture and Agri-Food Canada 227-BI Ozga Jocelyn University of Alberta 118-AB,

245-DV Pang Karlson University of British Columbia 144-CB Paudel Jamuna Risal Agriculture Agri-Food Canada 134-BS Peng Yujun University of British Columbia 127-BS Perkins Mendel University of British Columbia Pinhero Reena University of Guelph AP.5,

206-AP Pratiwi Putri Hokkaido University AB1.1 Pyc Michal University of Guelph CB1.3 Quilichini Teagen National Research Council, Saskatoon CDS.4 Rabbi Fazle Dept. of Chemistry and Biochemistry,

University of Regina 228-CW

Rahman Noabur University of Saskatchewan NM.3 Rahman Tawhidur University of Saskatchewan AB1.3,

248-DV Ravichandran Sridhar Agriculture and Agri-Food Canada AB2.5 Reinecke Dennis University of Alberta 243-DV Richardson Matt BCIT

128

Ro Dae-Kyun University of Calgary CDS.3 Romero Sergio University of Western Ontario Rosado Abel University of British Columbia CB1.6 Roveredo Jonathan Mount Royal University 226-BI Ruiz Orduna Alberto University of British Columbia 251-SM Samuel Marcus University of Calgary Samuels Lacey University of British Columbia Sarker Lukman UBC Okanagan Campus 255-SM Savada Raghavenrda

Prasad AAFC Saskatoon

Saxena Praveen University of Guelph AP.1 Scandola Sabine University of Calgary SR.3 Schiltroth Braeden Simon Fraser University 102-AB Schuetz Mathias Kwantlen Polytechnic University Selvaraj Kumarakurubaran Department of Biology, University of

Saskatchewan 157-SR

Shah Jay Terramera Shao Yang McGill University 112-AB Shoker Shamin University of British Columbia Singer Stacy AAFC Singh Santokh University of British Columbia 250-ED Skori Logan The University of Calgary DV.6 Sola Kresimir University of British Columbia SS.6 Song Changzheng University of British Columbia 103-AB Soolanayakanahally Raju Agriculture and Agri-Food Canada,

Saskatoon Research and Development Centre

SM2.6

Stanic Matija University of Calgary 108-AB Stobbs Jarvis Canadian Light Source TI.3 Stone Sophia Dalhousie University CDN.1 Sun Jiaqi McGill University CB1.5 Sun Menghan University of Manitoba 242-DV Sun Tongjun University of British Columbia BS.6 Sun Yulin University of British Columbia 213-BC Tai Helen Agriculture and Agri-Food Canada

Fredericton Research and Development Centre

BS.1

Tanino Karen University of Saskatchewan Tasnim Sifat Simon Fraser University SM1.3 Toth Ryan University of Calgary

129

Tran Anh University of Ottawa 236-DV Tran Lan University of British Columbia 230-CW Tremblay Jonathan Université Laval 214-BC Tripathi Rajiv Kumar McGill University 239-DV Unda Faride University of British Columbia 140-BS Utomo Joseph University of Calgary 254-SM Venglat Prakash University of Saskatchewan EP.6 Vishwanathan Kishore Georg-August-Universität, Göttingen,

Germany 221-BI

Wang Hong Dept. of Biochemistry, Univ. of Saskatchewan

SR.1

Wang Junfang University of British Columbia Wang Shucai Northeast Normal University AB2.2 Wang Shumin University of British Columbia 229-CW Wasteneys Geoffrey University of British Columbia Watanabe Yoichiro University of British Columbia CB2.5 Weismann Christina University of British Columbia 219-BI Wenzel Carol BCIT Wersch Rowan University of British Columbia BS.2 Whitehill Justin Michael Smith Laboratories DV.3 Willick Ian University of Saskatchewan 117-AB Wilson Ken University of Saskatchewan Wong Christopher University of Toronto Mississauga 211-BC Woodley Marcus University of British Columbia 146-CB Wu Di University of British Columbia 132-BS Xiong Yuanyuan University of British Columbia Xu Yang University of Alberta SS.5 Xu Yanqun Agriculture and Agri-Food Cananda Yang Xuan University of Western Ontario 231-CW Yao Danyu University of British Columbia EP.5 Yap-Dejeto Leni University of the Philippines, UC SC 199-AP Zhao Lifang University of British Columbia Zheng Hugo McGill University Zhou Chen Simon Fraser University 216-BT Ziegler Dylan Thompson Rivers University SR.6

130

Key AB Abiotic Stress AP Applied Plant Biology BC Biochemistry BI Biotic Interactions BS Biotic Stress BT Biotechnology/Tech Innovations CB Cell Biology CDN CD Nelson Lecture CDS Carl Douglas Symposium DV Development EP Ecophysiology NM Nutrients and Metabolism PLEN Plenary Session SM Specialized Metabolism SR Sexual Reproduction SS Seeds to Sky TI Technological Innovations

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