Abstract
Chromatin remodeling is important for the localization and function of many proteins involved in transcription, DNA replication and repair. Regulation of such activity within the chromatin context requires posttranslational modifications (PTM) of histones, such as ubiquitination, acetylation, methylation, sumoylation and phosphorylation that counterbalance the repressive nature of chromatin. In mammals, the FACT complex is essential for transcription initiation, relaxing chromatin condensation through displacement of the nucleosome. Being crucial in chromatin remodeling during transcription, and considering its general function as a chromatin remodeling facilitator, here we investigate the potential role of FACT during DNA repair mediated by homologous recombination (HR). In accordance with other previous works, our results also show that during transcription, FACT and RNF20, an E3 ubiquitin ligase recent found by our lab and others to be involved in DNA repair, interact with themselves. Moreover, this interaction seems to take place also during DNA damage response, which appears not to be related to transcription. Furthermore, depletion of FACT led to compromised DNA end resection through RAD51 and BRCA1 recruitment, whereas H2AX phosphorylation showed no major change. Additionally, RNF20 recruitment to the damage site and subsequent H2B ubiquitination, an important marker of chromatin relaxation, were also suppressed. Therefore, our data suggest that FACT might play a deciding function in chromatin assessment of other DNA repair proteins following DNA damage by loading of RNF20 to the damage sites. Together, they might play a critical role in HR independently of H2AX, revealing an alternative pathway of the chromatin remodeling during repair.
