Download Tissue specific hormone response and epigenome

Tissue specific hormone response and
epigenome reprogramming
Silin Zhong
State Key Laboratory of Agrobiotechnology
School of Life Sciences
The Chinese University of Hong Kong
Synopsis
• Use tomato fruit development and ethylene as a model
• Ethylene responsive genes
• Ethylene transcription factor binding
• Epigenetic (DNA methylation) changes
Ethylene
The McMurchie hypothesis
Repress ethylene
synthesis
System 1, self-inhibitory
Promote ethylene
synthesis
System 2, auto-catalytic
McMurchie et al. (1972) Nature; Theologis (1992) Cell;
Klee & Giovannoni (2011) Annu Rev Genet.
Transcription factor EIN3 is a master
regulator of ethylene signaling
C2H4
Ethylene
receptors
CTR
EIN2
EBF1/2
EIN3
activation of ethylene responsive genes
ERFs
chitinase
E4
PG
Chromatin immunoprecipiation (ChIP-Seq)
Crosslink transcription
factor to DNA
Use specific antibody to
capture TF-DNA complex
Recover antibody-TF-DNA
Sequence the DNA
Model-based analysis
Zhang et al., (2008) Genome Biol
DNA Methylation
EIN3 binding site at the promoter of E4
ChIP-Seq peak
Identify the EIN3 binding sites
Previous prediction based on EMSA
AYGWAYCT (Y=C/T, W=A/T)
Ethylene in fruit ripening
Grow
Ripening
Ethylene
Seed development
?
Use bisulfite sequencing to quantify
DNA methylation
Bisulfite converts C to U
After PCR, U becomes T
Sodium bisulfite
oxidative deamination
T/C mismatch = unmethylated C
C/C match = methylated C
Genome-wide demethylation during fruit ripening
Genebody
DNA methylation also changes near RIN
binding sites
RIN binding sites
called by ChIP-seq
Methylation influence TF binding?
TF
Tissue A
Tissue B
TF
Transcription
factor
Growth
hormone
Use RNA-Seq to identify ethylene responsive
genes
0 10 20 30 40 50 60 120
RNA-Seq
Gene expression profiles
Hormone response genes are different in
3 tissues
up-regulation
down-regulation
Transcription factor tissue-specific
binding site
Imature fruit tissue
Matured fruit tissue
Tissue specific binding site
Summary
• Use RNA-Seq to find hormone responsive genes
• Use ChIP-Seq to find transcription factor binding sites
• Use BS-Seq to quantify DNA cytosine methylation
• Differential transcription factor binding could be responsible
for tissue-specific hormone response
• In some promoters, DNA methylation changes are associated
with tissue-specific transcription factor binding sites
guilt by association is not a proof for causation
Maybe DNA methylation is a passive mark?
DME
TF
DME
TF
-
X
DNA demethylase (DME) could be the key
There could be another missing factor X
Acknowledgements
Jim Giovannoni
Leon Kochian
Zhangjun Fei
Jeffry Doyle
Jenny Xiang
Chris Watkins
Zongrang Liu
Eric Richard
Xiping Wang
Joe Ecker
Yong Xu
Bao Liu
Biyan Zhou
It is getting complicated
• Histone modification could also play a role
Active chromatin mark
Repressive mark