A novel variant of DNA polymerase ζ, Rev3 ΔC, highlights differential regulation of Pol32 as a subunit of polymerase δ versus ζ in Saccharomyces cerevisiae

Unrepaired DNA lesions often stall replicative DNA polymerases and are bypassed by translesion synthesis (TLS) to prevent replication fork collapse. Mechanisms of TLS are lesion- and species-specific, with a prominent role of specialized DNA polymerases with relaxed active sites. After nucleotide(s) are incorporated across from the altered base(s), the aberrant primer termini are typically extended by DNA polymerase zeta (pol zeta). As a result, pol zeta is responsible for most DNA damage-induced mutations. The mechanisms of sequential DNA polymerase switches in vivo remain unclear. The major replicative DNA polymerase delta (pol delta) shares two accessory subunits, called Pol31/Pol32 in yeast, with pol zeta. Inclusion of Pol31/Pol32 in the pol delta/pol zeta holoenzymes requires a [4Fe-4S] cluster in C-termini of the catalytic subunits. Disruption of this cluster in Pol zeta or deletion of POL32 attenuates induced mutagenesis. Here we describe a novel mutation affecting the catalytic subunit of pol zeta, rev3 Delta C, which provides insight into the regulation of pol switches. Strains with Rev3 Delta C, lacking the entire C-terminal domain and therefore the platform for Pol31/Pol32 binding, are partially proficient in Pol32-dependent UV-induced mutagenesis. This suggests an additional role of Pol32 in TLS, beyond being a pol zeta subunit, related to pol delta. In search for members of this regulatory pathway, we examined the effects of Maintenance of Genome Stability 1 (Mgs1) protein on mutagenesis in the absence of Rev3-Pol31/Pol32 interaction. Mgs1 may compete with Pol32 for binding to PCNA. Mgs1 overproduction suppresses induced mutagenesis, but had no effect on UV-mutagenesis in the rev3 Delta C strain, suggesting that Mgs1 exerts its inhibitory effect by acting specifically on Pol32 bound to pol zeta.The evidence for differential regulation of Pol32 in pol 8 and pol zeta emphasizes the complexity of polymerase switches. (C) 2014 Elsevier B.V. All rights reserved.