Deciphering the chromatin landscape induced around DNA double strand breaks

Cell Cycle. 2010 Aug 1;9(15):2963-72. doi: 10.4161/cc.9.15.12412.

Abstract

DNA double strand breaks (DSBs) are among the most deleterious forms of lesions and deciphering the details of the chromatin landscape induced around DSBs represents a great challenge for molecular biologists. Chromatin Immunoprecipitation, followed by microarray hybridisation (ChIP-chip) or high-throughput sequencing (ChIP-seq), are powerful techniques that provide high-resolution maps of protein-genome interactions. However, applying these techniques to study chromatin changes induced around DSBs was previously hindered due to a lack of suitable DSB induction techniques. We have recently developed an experimental system utilizing a restriction enzyme fused to a modified oestrogen receptor ligand binding domain (AsiSI-ER), which generates multiple, sequence-specific and unambiguously positioned DSBs across the genome upon induction with 4-hydroxytamoxifen (4OHT).(1) Cell lines expressing this construct represent a powerful tool to study specific chromatin changes during DSB repair, enabling high-resolution profiling of DNA repair complexes and chromatin modifications induced around DSBs. Using this system, we have recently produced the first map of gammaH2AX, a DSB-induced chromatin modification, on two human chromosomes and have investigated its spreading properties.(1) Here we provide additional data characterizing the cell lines, present a genome-wide profile of gammaH2AX obtained by ChIP-seq, and discuss the potential of our system towards investigations of previously uncharacterized aspects of DSB repair.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Base Sequence
  • Cell Cycle / drug effects
  • Cell Line, Tumor
  • Chromatin / metabolism*
  • Chromatin Immunoprecipitation
  • DNA Breaks, Double-Stranded* / drug effects
  • Fluorescent Antibody Technique
  • Genome, Human / genetics
  • Histones / metabolism
  • Humans
  • Nuclear Proteins / metabolism
  • Tamoxifen / analogs & derivatives
  • Tamoxifen / pharmacology

Substances

  • Chromatin
  • H2AX protein, human
  • Histones
  • Nuclear Proteins
  • Tamoxifen
  • afimoxifene