Two independent DNA repair pathways cause mutagenesis in template switching deficient Saccharomyces cerevisiae.

This article investigates two distinct mutagenic DNA repair pathways that operate in Saccharomyces cerevisiae when error-free template switching, mediated by Rad5-dependent PCNA polyubiquitination, is compromised. The study demonstrates that both Rad5-mediated and canonical PCNA monoubiquitination (PCNA-Ub)-mediated pathways recruit the translesion synthesis (TLS) DNA polymerase ζ via the scaffold protein Rev1, but differ in their modes of Rev1 recruitment and are genetically separable. Both pathways contribute additively to mutagenesis and replication stress response across early- and late-replicating genomic regions, with defects in either pathway leading to accumulation of single-stranded DNA and checkpoint activation. Furthermore, a natural RAD5 variant found in a wild yeast strain impairs PCNA polyubiquitination and error-free repair, thereby enhancing mutagenic repair and contributing to natural variation in mutation rates. These findings highlight the dual roles of Rad5 and PCNA modifications in regulating spontaneous mutagenesis and genome stability through distinct postreplication repair mechanisms.