The repair of abundant oxidative DNA damage is primarily performed by the Base Excision Repair (BER). BER of 8-oxoG in mammalian cells is initiated by the 8-oxoG-glycosylase 1 (OGG1). In the end of XXth century, the different steps of BER were studied in details on short pieces of naked DNA substrate. However, how BER operates on more physiological substrates such as chromatin was still unclear. To address this challenge, I established new tools and studied the mechanisms of BER by introducing a single 8-oxoG lesion in the DNA of reconstituted positioned nucleosomes using either conventional histone H2A or a histone variant H2A.Bbd, giving a more open structure to the nucleosome. BER was strongly reduced in both types of nucleosomes. An ATP-dependent chromatin remodeler, (e.g. SWI/SNF), was required to stimulate the BER. Our work disclosed that initiation of BER by glycosylases cannot overcome the nucleosomal barrier and requires specific chromatin remodeling.

In order to further understand BER mechanisms, I set up a novel technique allowing the specific formation of 8-oxoG in various cellular compartments. This novel live-cell imaging approach revealed a strong and very rapid recruitment of two NER-initiating factors (XPC and CSB) to sites of 8-oxoG in living cells. Interestingly, CSB exhibited different and transcription-dependent kinetics in the two compartments studied (nucleolus and nucleoplasm), suggesting a direct transcription-dependent involvement of CSB in the repair of oxidative lesions associated with different RNA polymerases but not involving other NER proteins. Based on our latest results, we propose a model in which CSB plays a role in facilitating BER progression at transcribed genes, probably to allow XRCC1 (an essential downstream BER factor) recruitment to BER-intermediates masked by RNA polymerase II complexes stalled at these intermediates.

Contact : karin.sadoul@univ-grenoble-alpes.fr