Transcript Mapping the DNA Damage Response

Mapping the DNA Damage
Response
Case study reveals
transcription factor (TF)
modules, dynamic TF
binding and an expanded
role for cell cycle regulators
Overview
• Experimental factors and selection
– Multiple criteria used
• ChIP-on-chip
– Differential binding analysis
• Gene expression of TF-deletion mutants
– Clustering analysis
– Deletion-buffering analysis
• Data integration and pathway reconstruction
Overview of the approach
Overview of the approach
Overview of the approach
Transcription factors that regulate
DNA damage response
Activated regulatory
network
Transcription factors that regulate
DNA damage response
Activated regulatory
network
TF knockout
“Deletion-buffered”
Growth
phenotype
in MMS:
mutants that
display relative
growth inhibition
Truncated Product Method (TPM):
determine condition dependent binding
ChIP-chip of 30 TFs before and
after DNA damage
YPD
MMS
+/-MMS
TPM
ChIP-chip Data
Summary
TFs may regulate different
genes (bind different
promoters) under different
conditions.
Workman CT, Mak HC, McCuine S, Tagne
JB, Agarwal M, Ozier O, Begley TJ, Samson
LD, Ideker T. A systems approach to
mapping DNA damage response
pathways. Science. 2006 May
19;312(5776):1054-9.
Promoter regions analysis
ChIP-chip and DNA-Motif
TF-Knockout expression profiles:
(look much like wild-type)
Environmental “epistasis analysis”:
(deletion-buffering)
Deletion-buffering analysis
Bayesian Score
Deletion-buffering examples
RNR Genes are repressed by
Rfx1p
Sensitive TFs are required for a greater
number of damage responsive genes
Integrated model
(regulatory paths explaining buffered genes)
Pathway
reconstruction
Pathway
reconstruction
Pathway
reconstruction
Integrated direct and
indirect regulatory
pathways (chIP-chip,
prot-prot) that
explain deletionbuffering
relationships
Workman CT, Mak HC, McCuine S, Tagne
JB, Agarwal M, Ozier O, Begley TJ, Samson
LD, Ideker T. A systems approach to
mapping DNA damage response
pathways. Science. 2006 May
19;312(5776):1054-9.
Summary
• “Sensitive” TFs control more of the DNA
damage response than non-sensitive TFs
• Regulatory networks are highly
interconnected
• Transcriptional regulation of important
DNA damage checkpoint kinases are
observed
• Measuring differential TF-binding is difficult