Two independent DNA repair pathways cause mutagenesis in template switching deficientSaccharomyces cerevisiae

Abstract

AbstractUpon DNA replication stress, cells utilize the post-replication repair pathway to repair single-stranded DNA and maintain genome integrity. Post-replication repair is divided into two branches: error-prone translesion synthesis, signaled by PCNA mono-ubiquitination, and error-free template switching, signaled by PCNA poly-ubiquitination. InSaccharomyces cerevisiae, Rad5 is involved in both branches of repair during DNA replication stress. When the PCNA poly-ubiquitination function of Rad5 is disrupted, Rad5 recruits translesion synthesis polymerases to stalled replication forks, resulting in mutagenic repair. Details of how mutagenic repair is carried out, as well as the relationship between Rad5-mediated mutagenic repair and the canonical PCNA-mediated mutagenic repair, remain to be understood. We find that Rad5-mediated mutagenic repair requires the translesion synthesis polymerase ζ but does not require other yeast translesion polymerase activities. Furthermore, we show that Rad5-mediated mutagenic repair is independent of PCNA binding by Rev1 and so is separable from canonical mutagenic repair. In the absence of error-free template switching, both modes of mutagenic repair contribute additively to replication stress response in a replication timing-independent manner. Cellular contexts where error-free template switching is compromised are not simply laboratory phenomena, as we find that a natural variant inRAD5is defective in PCNA poly-ubiquitination and therefore defective in error-free repair, resulting in Rad5- and PCNA-mediated mutagenic repair. Our results highlight the importance of Rad5 in regulating spontaneous mutagenesis and genetic diversity inS. cerevisiaethrough different modes of post-replication repair.