Pol32 is required for Pol zeta-dependent translesion synthesis and prevents double-strand breaks at the replication fork.
POL32 encodes a non-essential subunit of Poldelta and plays a role in Poldelta processivity and DNA repair. In order to understand how Pol32 is involved in these processes, we performed extensive genetic analysis and demonstrated that POL32 is required for Polzeta-mediated translesion synthesis, but not for Poleta-mediated activity. Unlike Polzeta, inactivation of Pol32 does not result in decreased spontaneous mutagenesis, nor does it limit genome instability in the absence of the error-free postreplication repair pathway. In contrast, inactivation of Pol32 results in an increased rate of replication slippage and recombination. A genome-wide synthetic lethal screen revealed that in the absence of Pol32, homologous recombination repair and cell cycle checkpoints play crucial roles in maintaining cell survival and growth. These results are consistent with a model in which Pol32 functions as a coupling factor to facilitate a switch from replication to translesion synthesis when Poldelta encounters replication-blocking lesions. When Pol32 is absent, the S-phase checkpoint complex Mrc1-Tof1 becomes crucial to stabilize the stalled replication fork and recruit Top3 and Sgs1. Lack of any of the above activities will cause double strand breaks at or near the replication fork that require recombination as well as Rad9 for cell survival.
Hanna
Michelle
M
Department of Microbiology and Immunology, University of Saskatchewan, 107 Wiggins Road, Saskatoon, Sask., Canada.
Ball
Lindsay G
LG
Tong
Amy H
AH
Boone
Charles
C
Xiao
Wei
W
D053903:DNA Breaks, Double-Stranded; D004265:DNA Helicases / Q000235:genetics / Q000378:metabolism; D004260:DNA Repair / Q000502:physiology*; D004261:DNA Replication; D004271:DNA, Fungal / Q000235:genetics / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D008954:Models, Biological; D016296:Mutagenesis; D009154:Mutation; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism / Q000528:radiation effects; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D014466:Ultraviolet Rays
D004271:DNA, Fungal; C522758:Pol32 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C061376:SRS2 protein, S cerevisiae; C425929:DNA polymerase zeta; D004259:DNA-Directed DNA Polymerase; D004265:DNA Helicases