Importance of microRNA for seedling

development in Arabidopsis thaliana

Jennifer CoppersmithDepartment of Biochemistry and BiophysicsMentor: Dr. Hiro Nonogaki (Horticulture)

Significance of Seed Development

pgrc3.agr.ca/images/seeds.gif

Agriculture and Food

Why Arabidopsis?

Arabidopsis Seed Structure

Embryo

Endosperm

Testa

Overview of Experiment

T DNA insert

SQUAMOSA PROMOTER-BINDING PROTEIN-LIKE13 (mSPL13)

SPL13

microRNA

• 21-24nt in length

• Gene regulation at the mRNA level

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RISCRISC

miRNA

microRNA

mRNA

miRNA

RISC

miRNA

microRNA mutation

mRNA

Mutation

Hypothesis: SPL13 needs to be down regulated by microRNA156 to maintain normal developmental program for germination or seedling establishment.

GUGCUCUCUCUCUUCUGUCA CACGAGUGAGAGAAGACAGU

GUGCUCUCUCUCUUCUGUCA CACGAGUGAGAGAAGACAGU

A A A A

SPL13

miR156

mSPL13

WT Cont mSPL13

rRNA

SPL13

Overaccumulation of SPL13

Mutant Phenotype

Cot

Cot

4 leaf seedling (normal) 2 leaf seedling (delayed)

Cot

Cot

2-leaf seedlings4-leaf seedlings

Developmental Delay

Temporal Seedling Arrest

WTmSPL13

CONTROL

0 d

Shoot Apical Meristem (SAM)

2 d

Storage Protein and Meristem

Meristem

Embryo

Cot

Hypocotyl

Radical

WT

1 d 3 d2 d 4 d

mSPL13

mSPL13WT

3 d

mSPL13WT

2 d

mSPL13WT

4 dMeristem Development

mSPL13WT

8 d

Visible Difference

Analysis

WT mSPL13

SPL13

Based on phenotype, down regulation of SPL13 via microRNA is essential for proper meristem

development.

Determine Potential Targets of SPL13

miRNA156

SPL13 SPL13

Promoter Gene X

X

AcknowledgementsHiro Nonogaki

Kevin Ahern

Howard Hughes Medical Institute

Lab Members:

Masa Asahina

Tammy Chan

Po-Pu Liu

Jessica Kristof

Piotr Pupel

Jing Sun

SQUA

SQUAMOSA ( or APETALA1)

10 bp conserved stretch

19-mer core seq

- 419 - 393

- 455 - 429

AP1

T T C G A C G T C C G T A C A A C A A G T C T T T A T

C C A G T G G T C C G T A C A A T G T T A C T T A C C

SBP1 SBP2SPL13

?

Normal Embryogenesis in mSPL13

Coppersmith JL, Hewitt JR, Nonogaki H, unpublished results