The majority of environmental stressors act, at least in part, via the induction of damage to DNA, triggering genomic instability, which is linked to a variety of cancers. Presently, the associations between damage and disease are based upon rather crude assessments of total genome levels of damage, which provide little mechanistic insight. Increasingly, it is clear that genomic location, and persistence of damage, is of greater biological significance, as this may lead to aberrant outcomes, such as mutation; promotion of microsatellite instability; modulation of gene expression through affecting transcription factor binding, and DNA methylation; and acceleration of telomere shortening. Such a diverse range of effects may explain why such damage has a role in both malignant and non-malignant disease. In order to understand the link between DNA damage and downstream cellular (and organismal) consequences, we need informative methods for assessing the location of DNA damage, which take into account genomic organization. In response, reports are now beginning to emerge which describe genome-wide approaches for mapping DNA, some of which have nucleotide resolution. We have developed a method which combines immunoprecipitation of damage-containing DNA, with next generation sequencing (which we have termed DDIP-seq). Using this approach we have mapped the formation and repair of solar simulated radiation-induced DNA damage (cyclobutane thymine dimers; T<>T) across the entire genome, in human skin keratinocytes. We identified a profound differential distribution in both the formation and repair of T<>T, across the entire genome. Key findings included: induced levels of T<>T vary significantly across the genome, indicating the existence of regions of sensitivity (and conversely resistance) to formation. Furthermore, some genomic regions are fully repaired within 24 h, whereas a significant number of regions retain >90% of damage 24 h after exposure. This contrasts with the assessment of global genome repair of T<>T, estimates of which range from 50% of T<>T remaining at 24 h and ∼40-47% remaining at 48 h, and illustrate the extent to which total DNA damage/repair measures fail to reflect events in discrete sequences. These results further demonstrate that the rate of DNA repair at a specific locus is independent of the initial level of damage. We are now combining these data with parallel ChIP-seq analysis of chromatin remodeling, to take into account genomic organization. Our findings stress the importance of genome-wide, sequence specific approaches and the more detailed information that they provide. to help us better understand the link between DNA damage and cancer.

Citation Format: Alaa Alhegaili, George D. Jones, Marcus S. Cooke. Genome-wide analysis of DNA damage and repair reveals differential sites and rates of repair, together with differential sensitivities to damage. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-163.