Defective interaction between Pol2p and Dpb2p, subunits of DNA polymerase epsilon, contributes to a mutator phenotype in Saccharomyces cerevisiae

Abstract

SummaryMost of the prokaryotic and eukaryotic replicative polymerases are multi-subunit complexes. There are several examples indicating that noncatalytic subunits of DNA polymerases may function as fidelity factors during replication process. In this work, we have further investigated the role of Dpb2p, a noncatalytic subunit of DNA polymerase epsilon holoenzyme from Saccharomyces cerevisiae in controlling the level of spontaneous mutagenesis. The data presented indicate that impaired interaction between catalytic Pol2p subunit and Dpb2p is responsible for the observed mutator phenotype in S. cerevisiae strains carrying different mutated alleles of the DPB2 gene. We observed a significant correlation between the decreased level of interaction between different mutated forms of Dpb2p towards a wild-type form of Pol2p and the strength of mutator phenotype that they confer. We propose that structural integrity of the Pol epsilon holoenzyme is essential for genetic stability in S. cerevisiae cells. KeywordsDNA polymerase; Pol epsilon holoenzyme (Pol ε HE); Spontaneous mutagenesis; Fidelity of DNA replication; Protein-protein interaction; DPB2 (YPR175W) and POL2 (YNL262W) genes of the yeast Saccharomyces cerevisiae

Fidelity consequences of the impaired interaction between DNA polymerase epsilon and the GINS complex

“…It has been established that the C-terminus of Dpb2p is important for its interaction with Pol2p [49]. Isolated in our laboratory dpb2 mutants exhibit temperature-sensitivity and mutator phenotype [47,48]. We observed a strong synergistic mutator effect between the mutant alleles of DPB2 and msh6 mutation suggesting that dpb2-dependent errors arising during DNA replication are substrates for mismatch repair system.…”

“…We have previously isolated several temperature-sensitive mutant alleles of DPB2 encoding the non-catalytic subunit of Pol holoenzyme in the yeast S. cerevisiae [47,48]. The mutants were significantly affected in spontaneous mutagenesis.…”

“…A few years ago we found that Dpb2p, a non-catalytic subunit of DNA Pol , is important for faithful replication of genomic DNA [47,48]. It has been established that the C-terminus of Dpb2p is important for its interaction with Pol2p [49].…”

DNA Polymerase ε

“…Temperature sensitive dpb2 alleles were shown to give an increased spontaneous mutation rate (Jaszczur et al 2008 ) . The level of spontaneous mutation rates were correlated with loss of interaction with Pol2 in two-hybrid assays (Jaszczur et al 2009 ) . Whether Dpb2 directly in fl uences the fi delity of Pol2 remains to be shown.…”

“…These interact with a motif closer to the N-terminus of Pol2 (Shikata et al 2006 ) . Yeast two-hybrid assays have also shown that yeast Dpb2 interacts with the zinc-fi nger domain (Dua et al 1998 ;Jaszczur et al 2009 ) . It is entirely possible, therefore, that the very C-terminal domain of Pol2 extends away from the globular head domain and provides binding sites for the accessory subunits.…”