mapping the dna damage response

Case study reveals transcription factor (TF) modules, dynamic TF binding and an expanded role for cell cycle regulators

Mapping the DNA Damage Response

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

TF knockout

“Deletion-buffered”

Activated regulatory network

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

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.

TFs may regulate different genes (bind different promoters) under different conditions.

Promoter regions analysisChIP-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 deletion-buffering 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