PRJNA449230
Rad5 recruits TLS DNA polymerases in replication stress to repair ssDNA gaps on undamaged templates
Post-replication repair (PRR) allows tolerance of chemical- or UV-induced DNA lesions in both an error-free and an error-prone manner. In classical PRR, PCNA monoubiquitination recruits translesion synthesis (TLS) DNA polymerases that can replicate through lesions. Polyubiquitination of PCNA by the ubiquitin ligase Rad5 initiates error-free PRR, using the sister chromatid as a template for DNA synthesis in lieu of the lesion-containing strand. We find that Rad5 forms nuclear foci during normal S-phase and after exposure to types of replication stress where DNA base lesions are likely absent. We wanted to identify what Rad5 foci correspond to in cells, what signals for Rad5 relocalization and what the role of Rad5 is in the replication stress response. Using Chromatin immunoprecipitation followed by deep sequencing (ChIP-seq) we demonstrate that after dNTP depletion leading to replication fork stalling Rad5 is enriched on chromatin in a similar pattern to the replicative polymerases , indicating that Rad5 binds stressed-replication forks. Rad5 binds and recruits Rev1 to stressed replication forks to mediate repair ssDNA and prevent mitotic defects and chromosome breaks. Our data indicate that Rad5 is the central effector of PRR signaling at stressed replication forks, where Rad5 promotes mutagenic repair of undamaged ssDNA that arises during physiological and exogenous replication stress.