POSTERS

Poster 1

Drought stress experiments in tropical trees

Aneth Sarmiento and Margarete Baier

Dahlem Centre of Plant Sciences, Institute of Plant Physiology, Freie Universität Berlin,

Berlin, Germany

The response of tropical plants to stress has been barely investigated. Due to the

climate change, drought is one of the major problems. This situation could have

consequences for photosynthesis rate. Under drought stress, C3 plants are known to

close stomata and to stop photosynthesis. This leads to an accumulation of electrons

and an increase in ROS (Reactive Oxygen Species) production. To avoid their

negative effects, a strong antioxidant system is required.

We have evaluated three tropical plants that have different photosynthetic pathways

and differ in the tolerance to drought. Swietenia macrophylla (C3, drought sensitive),

Jatropha curcas (C3, probably switching to CAM, drought tolerant) and Clusia rosea

(CAM), from Panama, were grow in a greenhouse under similar conditions as in

nature.

As the plants under investigation are not sequenced, at the moment, we work on

each individual sequence. Partial sequence of 2-cys- peroxiredoxin (2CP) and

ascorbate peroxidase (APX) were discovered in the lab after a phylogenetic study

based on amino-acid similarities, to study gene expression regulation during stress.

We expect soon to perform stress experiment to measure gene expression.

POSTERS

Poster 2

The cytokinin status primes light-dependent seed germination in

Arabidopsis thaliana

Daniela Pezzetta, Stefanie Zintl, Jan-Erik Leuendorf, Michael Riefler, Thomas Schmülling

Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany

Seed germination is of crucial importance for a plant’s life history. In Arabidopsis

thaliana, seed germination is controlled by external cues, in particular light, and

endogenous factors such as hormones. However, the functional role of the

phytohormone cytokinin (CK) in germination has been investigated rarely.

Here we show that A. thaliana seeds with a lowered cytokinin content and seeds

impaired in the perception of cytokinin have a dramatically increased light sensitivity

of germination as compared to wild type, especially in the phytochrome A (phyA)-

dependent ‘very low fluence response’ (VLFR).

The cytokinin status may act as a priming stimulus altering the germination

phenotype of the progeny after a triggering light stimulus. We could show that

reduced CK perception of maternally derived seed tissue is particularly relevant to

enhance germination under VLFR conditions. Investigation of eventual

transgenerational priming effects of parental plants on their offspring revealed only a

minor influence of the parental light environment but a major impact of the parental

nitrogen availability on germination under VLFR conditions.

POSTERS

Poster 3

Plastids alter leaf morphology in plastome-genome-incompatible

Oenothera hybrids

Elena Ulbricht-Jones, Mark Aurel Schöttler, Patrick Giavalisco, Stephan Greiner

Max Planck Institute of Molecular Plant Physiology, Potsdam-Golm, Germany

The plant genome comprises the nuclear genome, the plastid genome (plastome)

and the mitochondrial genome. Nuclear-organellar interaction is crucial for plant

development and function, consequently resulting in coevolution of the genetic

compartments. Hence, new combinations of nuclear and cytoplasmic genomes can

cause incompatibilities even between closely related species.

An interesting plastome-genome-incompatibility altering leaf morphology can be

observed in certain interspecific Oenothera hybrids. Reciprocal crosses between

Oenothera berteriana and O. odorata yield hybrids which have identical nuclear

genomes combined with differing cytoplasms. Hybrids hosting O. berteriana plastids

have green and broad leaves, whereas O. odorata plastids are incompatible with the

hybrid nuclear genome and confer pale and narrow leaves. We uncovered 146

polymorphisms distinguishing O. odorata from O. berteriana plastomes, representing

a broad range of loci potentially responsible for the observed leaf phenotype. An

association mapping approach based on full plastome sequences of various

Oenothera accessions helped narrow down the list to 46 putative candidates. We

could exclude most of them by complementary molecular analyses, which rather

point towards lipid biosynthesis to be involved in the plastome-genome-

incompatibility and altered leaf morphology.

POSTERS

Poster 4

Comparative studies of plant latex proteins from Euphorbia

coerulescens HAW and Euphorbia fortissima L.C.LEACH with hen

egg white lysozyme

Florian Günther, Matthias F. Melzig

Dahlem Centre of Plant Sciences, Institute of Pharmacy - Pharmaceutical Biology, Freie

Universität Berlin, Berlin, Germany

Lysozyme is an enzyme from the group of hydrolases classified in the enzyme

classification group EC 3.2.1.17. It was discovered by A. Fleming in 1922 as an

element of the innate immune system and serves as a defense against Gram-positive

and Gram-negative bacteria. It acts as a mucolytic and is present in many body fluids

such as sweat, saliva and nasal secretions, as well as hen egg white, milk and

expressed vegetables [1, 2]. It is known that latices of the plant family Euphorbiaceae

Juss. show lysozyme activity [3]. In this study, latices of Euphorbia coerulescens

Haw. and Euphorbia fortissima L.C.Leach were tested by MALDI-TOF-MS in terms of

their sequence coverage with hen egg white lysozyme (HEWL). For this purpose

relevant samples were separated by SDS-PAGE electrophoresis; the relevant protein

bands were excised and processed using tryptic-in-gel-digestion. As a positive

control, HEWL was treated like the latex samples. The results show that the 33 kDa

bands in the samples of E. fortissima have sequence coverage with HEWL of 36.7%

and from E. coerulescens of 23,8%. Results with a sequence accordance greater

than 20% and at least two major peptides detected are to be regarded as a

significant positive [4]. By that it can be concluded that lysozyme is present in latices

of Euphorbiaceae and seems to be a highly conserved protein comparable with

isoenzymes in animals.

[1] Wang S. et al. Isolation and identification of a plant lysozyme from Momordica charantia

L.. Eur. Food Res Technol (2011) 613-619

[2] Meyer K. et al. Lysozym of plant origin. J. Biol. Chem. (1946) 733-740

[3] Guenther F. Investigation of the latices of Euphorbiaceae - genus Euphorbia - in terms of

lysozyme and chitinase activity. Ernst-Moritz-Arndt-Universität Greifswald, Diplomarbeit

(2013)

[4] Ali M. et al. Characterization and modeling oft the interactions between coffee storage

proteins and phenolic compounds. Journal of Agricultural and food Chemistry (2012) 11601-

11608

POSTERS

Poster 5

The plant´s transcriptome “forgets” a reliable stress signal when it

ceases, but “remembers” an unspecific past stress

when exposed to herbivory

Jana Oberländer1, Vivien Firtzlaff2, Sven Geiselhardt2, Monika Hilker2, Reinhard Kunze1

1Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany 2Dahlem Centre of Plant Sciences, Applied Zoology and Animal Ecology,

Freie Universität Berlin, Berlin, Germany

Biotic and abiotic stress responses of plants have been studied widely. However little

is known whether plants can remember a past stress and how herbivory-induced

transcriptome changes are affected by a previous stress. Using Arabidopsis thaliana

and the herbivore Pieris brassicae, we studied the effect of prior chilling or egg

deposition as a non-reliable or reliable herbivory indicating warning signal,

respectively. A five-day-chilling period induced ≥2-fold transcriptional changes of

5.417 genes. When these plants were retransferred to 20°C for one day and

subsequently exposed to herbivory for two days, 2.055 genes were differently

expressed compared to herbivory-exposed but chilling-inexperienced plants,

indicating a reprogramming of the plant's response to herbivory by a preceding

chilling stress. In contrast, the plant´s transcriptome did not show any memory of a

five-day-lasting egg deposition period one day after egg removal. Transcript levels of

all 647 egg-induced genes were reset to those of untreated control plants.

Accordingly, the plants “forgot” the reliable egg deposition-signal after its removal, but

“remembered” a past herbivore-unrelated chilling stress.

POSTERS

Poster 6

CPK5-mediated crosslink between SA- and ABA signaling

Jennifer Bortlik and Tina Romeis

Dahlem Centre of Plant Sciences, Institute of Biochemistry of Plants, Freie Universität Berlin,

Berlin, Germany

In nature, plants are continuously threatened by abiotic and biotic stresses.

Phytohormones are involved in signaling responses to environmental stresses but

also direct growth and development. In particular, crosstalk between different

hormone-signaling pathways has been implicated in the fine-tuning of plant defense

responses. For example, abscisic acid (ABA) controls abiotic stress responses and

plant development, but there is increasing evidence that ABA is also involved in

plant-pathogen interactions. In Arabidopsis the calcium-dependent protein kinase

(CDPK) isoform CPK5 is biochemically activated after pathogen-recognition.

Enhanced expression of salicylic acid (SA) marker genes as well as increased SA

hormone levels in CPK5 overexpression lines correlated with a SA-dependent

suppression of bacterial growth. Interestingly, treatment of plants with ABA reverts

the phenotypic as well as the molecular characteristics of enhanced CPK5 signaling,

accompanied by ABA-dependent changes in the CPK5 protein phosphorylation

pattern. We thus propose CPK5 as a potential hub in the crosslink between the SA

and ABA signaling pathways.

POSTERS

Poster 7

Long cis-natural antisense transcripts regulate the activity of

cytokinin metabolic genes and cytokinin homeostasis

in Arabidopsis

Jenny Engelman and Tomáš Werner

Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany

The phytohormone cytokinin is an important regulator of numerous physiological and

developmental processes in plants. For example, cytokinin controls cell division and

differentiation, and thus activities of plant meristems and the formation and growth of

organs during a plant’s life cycle. One important mechanism regulating cellular

cytokinin concentrations is the metabolic degradation catalyzed by cytokinin

oxidase/dehydrogenase (CKX) enzymes. Recently, we identified long naturally

occurring antisense transcripts at several CKX loci and started to investigate the

functional relevance of the NATs from one selected locus by reducing their activity in

amiRNA-expressing Arabidopsis plants. These plants show phenotypic and

molecular alterations that strongly resemble the cytokinin deficiency syndrome

caused by overexpression of CKX-coding transcripts. Indeed, the transgenic plants

have severely reduced cytokinin levels caused by an increased CKX activity.

Surprisingly, our analyses indicate that the investigated CKX NATs do not regulate

the cognate coding transcript in cis, but rather act negatively on phylogenetically

closely related CKX transcripts in trans.

POSTERS

Poster 8

Investigation of the triterpene-dependent uptake of mistletoe lectin

by adherent and suspension cells

Katharina Mulsow1, Thomas Enzlein2, Catharina Delebinski3, S. Jäger4, Georg Seifert3,

Matthias F. Melzig1

1Dahlem Centre of Plant Sciences, Institute of Pharmacy - Pharmaceutical Biology, Freie

Universität Berlin, Berlin, Germany 2Department of Biotechnology, Mannheim University of Applied Sciences, Mannheim,

Germany 3Department of Paediatrics, Division of Oncology ⁄ Haematology, Charité Universitätsmedizin

Berlin, Berlin, Germany 4 Birken AG, Niefern-Öschelbronn, Germany

Alternative treatments become more relevant due to the severe side effects of the

chemotherapeutics used in the first-line therapy. Viscum album L. (Santalaceae)

belongs to the phytotherapeutic drugs applied in cancer treatment [1]. The main

ingredients of the aqueous extracts are the mistletoe lectins which belong to the

ribosome-inactivating proteins type II [2]. The triterpenes, mainly oleanolic acid,

belong to the lipophilic components of the mistletoe.

In this work, the uptake of mistletoe lectin by THP 1-, HL 60-, 143B- and Ewing TC

71-cells were determined after 30, 60 and 120 minutes by an enzyme linked

immunosorbent assay. The isolated lectin and the aqueous extract, with or without

added mistletoe triterpenes, were analyzed.

After 120 minutes, an intracellular uptake of 20% was reached in all cell lines by

aqueous extract with added triterpenes. Furthermore, the 143B-cells - unlike the

other cell models - pick up mistletoe lectin only with concomitant mistletoe

triterpenes. The uptake in THP 1-, HL 60- and Ewing TC 71-cells was time-

dependent. The addition of triterpenes did not influence the process.

Interestingly, the uptake of mistletoe lectin by 143B-cells could only be measured

after addition of triterpenes. The reason of this special behavior is still not known and

needs further investigations.

[1] Grothey, A. et al. (1998) Dtsch Med Wochenschr 123: 923–9

[2] Olsnes, S. et al. (1982) J Biol Chem 257: 13263-70

POSTERS

Poster 9

The role of cytokinin N-glycosylation for cytokinin responses in

Arabidopsis thaliana

Louisa Brock and Tomáš Werner

Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany

Cytokinins are plant hormones which affect many aspects of plant growth and

development. Metabolic conjugation of cytokinins with sugars, forming O- and

N-glucosides, leads to inactivation. So far, two enzymes with cytokinin

N-glucosyltransferase activity, UGT76C1 and UGT76C2, have been identified in

Arabidopsis. To study the physiological role of these enzymes in planta we generated

gain- and loss-of-function mutants of the corresponding genes. In contrast to earlier

studies, strong overexpression of 35S:UGT76C2-GFP led to multiple phenotypic

changes that resembled those of cytokinin-deficient plants. The reduced cytokinin

status was supported by reporter gene studies and cytokinin response assays. As

single ugt76c1 or ugt76c2 mutants do not show obvious morphological changes, we

established artificial microRNA lines to silence both genes in parallel. Surprisingly,

simultaneous downregulation of UGT76C1 and UGT76C2 resulted in phenotypic and

molecular alterations that were, at least partially, similar to those of 35S:UGT76C2-

GFP-expressing plants.

POSTERS

Poster 10

Skyrocketing sales of medicinal products from Black Cohosh

(Actaea racemosa L.) – An answer to challenges in conservation of

the plant and adulteration of the herbal drug

Marian Bittner, R. Schenk, Matthias F. Melzig

Dahlem Centre of Plant Sciences, Institute of Pharmacy - Pharmaceutical Biology, Freie

Universität Berlin, Berlin, Germany

Dietary supplements and medicinal products from Actaea racemosa L.

(Ranunculaceae, Black Cohosh) are widely used to treat climacteric complaints as an

alternative to hormone substitution. Recent trials proof efficacy and safety of the

approved products made from extracts of pharmaceutical quality.

This led to increasing sales in the US (+350% from 2010 to 2014) on the one hand,

and on the other hand to increasing wild-harvests of A. racemosa rhizomes. Already

in 2010 about 143 t of dried plant material were aggregated. Today in some US

states the plant is classified as endangered. Furthermore the increasing demand for

the plant material leads to problems with accidental and intentional adulteration, but

the identity of the plant material is crucial for the drug’s safety.

The answer on all these challenges could be crop science and cultivation of

A. racemosa. But the homogeneity and quality of such plants derived by cultivation,

has never been investigated. Therefore we phytochemically characterized a large

number of 157 of such plants by validated RP-PDA-UPLC and HPTLC methods. We

could show the remarkable homogeneity of the plants and compared them to plants

from different origins from all over the world. The data supports the application of

cultivation for A. racemosa, which should be furthered.

POSTERS

Poster 11

Reil1-1 reil2-1 double mutant “in the grip” of cold stress:

Morphological and physiological phenotypes.

Olga Beine-Golovchuk and Joachim Kopka

Max Planck Institute of Molecular Plant Physiology, Potsdam-Golm, Germany

The Rei1 and Reh1 proteins are yeast paralogs that are part of the maturation

machinery of the 60S eukaryotic large ribosomal subunit. Deletion of Rei1 or of both

in yeast and Arabidopsis thaliana paralogs causes cold sensitivity. Reduced growth

of the deletion mutants at low temperature is associated with an aberrant polysome

profile that is characterized by accumulation of free 40S and 60S subunits. The reil1-

1 reil2-1 double mutant is even more cold-sensitive but, in contrast, has only a

marginal growth defect at optimal temperature. This surprising match of growth

defects between yeast and A. thaliana led us to investigate whether the function of

REIL genes is conserved and whether the reil genes may have acquired new

functions in a plant. In the presented work we focused on the characterization of the

reil1-1 reil2-1 mutant which is an extreme dwarf but still viable at constant 10°C. Our

data suggest that the reil1-1 reil2-1 has a growth defect that is linked to the

acclimation process to mild cold (10°C). In order to understand the underlying

mechanism for which the double mutant is deficient and the implied link to ribosomal

function we initiated an integrated systems analysis of short- and long-term,

transcriptomic and metabolomic responses to 10°C cold which we complement by

comparative analyses of ribosome composition and activity.

Stefanie Schmidt, Frederik Dethloff, Olga Beine-Golovchuk, Joachim Kopka (2013) The

REIL1 and REIL2 proteins of Arabidopsis thaliana are required for leaf growth in the cold.

Plant Physiology 163: 1623–1639 (DOI: 10.1104/pp.113.223925)

Schmidt S, Dethloff F, Beine-Golovchuk O, Kopka J (2014) REIL proteins of Arabidopsis

thaliana interact in yeast-2-hybrid assays with homologs of the yeast Rlp24, Rpl24A,

Rlp24B, Arx1 and Jjj1 proteins. Plant Signaling and Behavior 9: e28224 (DOI:

10.4161/psb.28224)

POSTERS

Poster 12

Epidermis-derived cytokinin regulates the growth and development

of shoot organs in Arabidopsis thaliana

Sören Werner, Ingmar Staude, Isabel Bartrina and Thomas Schmülling

Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany

Plants progress through a number of developmental transitions during their life cycle.

In the vegetative phase plants rapidly increase their size and mass, and in the

reproductive phase they produce flowers and seeds. The transitions from juvenile to

adult vegetative growth and further to flowering are regulated by a complex genetic

program that is influenced by both environmental and endogenous cues. The

phytohormone cytokinin (CK) plays a role in many aspects of plant growth and

development. It is known for instance that a change of the CK status in the plant can

influence the size and morphology of organs, as well as affect flowering time. The

regulatory mechanisms underlying these processes are still poorly understood. We

have generated transgenic Arabidopsis plants using an epidermis-specific promoter

in order to study the potential role of the outermost cell layer in regulating CK-

dependent developmental processes in shoot organs. Here we show that epidermis-

derived CK affects not only plant height and the size of lateral organs, but also

regulates the juvenile-to-adult phase transition, most probably by promoting miR172

expression in the plant.

POSTERS

Poster 13

Study of the complex formation between CKX and HIPP proteins

Tianqi Guo, Georgeta Leonte, Henriette Weber and Tomáš Werner

Dahlem Centre of Plant Sciences, Institute of Biology/Applied Genetics,

Freie Universität Berlin, Berlin, Germany

The concentration of biologically active cytokinins in the plant is effectively

downregulated through glycoconjugation and oxidative degradation catalyzed by

intra- and extracellular cytokinin oxidase/dehydrogenase (CKX) enzymes. In our

yeast two-hybrid studies, we identified several heavy metal-associated isoprenylated

plant proteins (HIPPs) as CKX-interacting proteins and confirmed the protein-protein

complex formation by co-immunoprecipitation experiments. In Arabidopsis, around 50

HIPP proteins can be defined by the plant-specific motif combination of one or two

heavy metal-binding domains (HMA) and a C-terminal isoprenylation motif. Very little

is known about the biological function of these proteins. The phenotypic and

molecular analysis of HIPP-overexpressing plants and hipp knock-out mutants

revealed changes in cytokinin status and responses in both types of plants

suggesting that the HIPP-CKX interaction is physiologically relevant. The possible

mechanisms underlying the HIPP-CKX interaction and how this might modulate the

activity of CKX proteins remain to be elucidated.

POSTERS

Poster 14

Medicinal Plants traditionally used for treatment of obesity and

Diabetes mellitus – Screening for

Pancreatic Lipase and α-Amylase inhibition

Tina Buchholz, Matthias F. Melzig

Dahlem Centre of Plant Sciences, Institute of Pharmacy - Pharmaceutical Biology, Freie

Universität Berlin, Berlin, Germany

Inhibition of digestive enzymes is one of the most widely studied mechanisms for the

treatment of obesity and its associated diseases as diabetes mellitus, coronary heart

diseases or sleep-breathing disorders.[1,2] The use of plant based resources as a

potential platform for discovery and development of new drugs has become a

lucrative research field in this context.

In order to find new pancreatic lipase (PL) and α-amylase inhibitors from natural

sources, 32 medicinal plants with weight-reducing, serum glucose-reducing or related

potential were investigated. Methanolic and water extracts of the

plants were evaluated by using two in vitro test systems. Our findings have shown

that the methanolic extract of Hibiscus sabdariffa L. (Malvaceae) showed the best

inhibitory activities to PL (IC50: 35.8 ± 0.8 µg/mL) and α-amylase (IC50:

29.3 ± 0.5 µg/mL). This works provides a priority list of interesting plants for further

study with respect to the treatment of obesity and associated diseases.

[1] Trigueros L et al. (2013). Food Ingredients as Anti-Obesity Agents: A Review. J. Food

Sci. Nutr. 53: 929-942.

[2] Foster-Schubert KE, Cummings DE (2006). Emerging Therapeutic Strategies for Obesity.

Endocr. Rev. 27: 779-793.