ChIP-Seq genome wide analysis of γH2AX induced by X-rays or heavy ions*

Heavy ions are extensively used at GSI as a tool to study DNA repair processes and chromatin structure dynamics. Ionizing radiation (IR) produces DNA double strand breaks (DSB) triggering the DNA damage response (DDR). One of the earliest events of DDR consists of the extensive phosphorylation of H2AX histone variant (γH2AX), whose distribution throughout the genome appears to be non-random. γH2AX spreads through megabases from the DNA DSB and its distribution has been reported to be uneven and asymmetric. However, in previous experiments DNA DSB had been induced by enzymes. Whilst this offers some advantages (e.g. position of DNA damage is known), on the other hand this method can be biased by efficiency and limiting factors (e.g. DNA damage induction cannot be quenched). Moreover, IR-induced DSB may be qualitatively different from those introduced enzymatically into the genome. Although it has been proposed that chromatin density as well as cellular metabolism (e.g. transcription) may regulate the γH2AX spreading, there is little evidence of such a mechanism on a genome scale. Next generation sequencing technologies offer the possibility to address issues on a genome scale. Coupling such high-throughput technology to chromatin immunoprecipitation (ChIP) provides a powerful tool (ChIP-Seq) to map the position of the desired target throughout the genome. We are using ChIP-Seq to provide a genome scale precise map of IR-induced γH2AX. Our goal is to describe how such histone modification propagates in the context of chromatin density.