Academic journal article Genetics

Mutagenic and Recombinagenic Responses to Defective DNA Polymerase [Delta] Are Facilitated by the Rev1 Protein in Pol3-T Mutants of Saccharomyces Cerevisiae

Academic journal article Genetics

Mutagenic and Recombinagenic Responses to Defective DNA Polymerase [Delta] Are Facilitated by the Rev1 Protein in Pol3-T Mutants of Saccharomyces Cerevisiae

Article excerpt

ABSTRACT

Defective DNA replication can result in substantial increases in the level of genome instability. In the yeast Saccharomyces cerevisiae, the pol3-t allele confers a defect in the catalytic subunit of replicative DNA polymerase δ that results in increased rates of mutagenesis, recombination, and chromosome loss, perhaps by increasing the rate of replicative polymerase failure. The translesion polymerases Pol η, Pol ζ, and Rev1 are part of a suite of factors in yeast that can act at sites of replicative polymerase failure. While mutants defective in the translesion polymerases alone displayed few defects, loss of Rev1 was found to suppress the increased rates of spontaneous mutation, recombination, and chromosome loss observed in pol3-t mutants. These results suggest that Rev1 may be involved in facilitating mutagenic and recombinagenic responses to the failure of Pol δ. Genome stability, therefore, may reflect a dynamic relationship between primary and auxiliary DNA polymerases.

THE cell has a large arsenal of mechanisms for preventing genome instability in the form of mutations, genome rearrangements, and loss of heterozygosity (LOH). Efficient DNA replication is critical for normal cellular function, not only because of the necessity to duplicate the genetic information, but also because faulty replication influences the spontaneous frequencies of mutation, genome rearrangement, and LOH arising from nicks, gaps, and breaks in DNA (Horiuchi et al. 1994; Ivessa et al. 2000; Saleh-Gohari et al. 2005). While a variety of DNA repair pathways, including homologous recombination, can provide an efficient and effective means of repairing such DNA damage (Michel et al. 2001; Garg and Burgers 2005a), without the appropriate controls they themselves may lead to increased genome instability (Petes and Hill 1988). These controls are critically important as elevated genome instability can lead to cell death, tumorigenesis, and the development of a range of complex diseases in humans. The normal function of systems involved in DNA replication, recombination, and repair are crucial as they have interdependent responsibilities in maintaining genomic integrity.

DNA replication in yeast is catalyzed by the primary replicative polymerases α, δ, and ε. Pol α synthesizes the primers for leading and lagging strand synthesis, while Pol δ and Pol ε are responsible for the bulk of bidirectional DNA replication (Garg and Burgers 2005b; Johnson and O'Donnell 2005; Pursell et al. 2007; NickMcElhinny et al. 2008). Strains carryingmutations in the POL1, POL2 (CDC17), and POL3 (CDC2) genes, which encode the catalytic subunits of polymerases α (Budd and Campbell 1987), ε (Boulet et al. 1989), and δ (Morrison et al. 1990), respectively, display increased rates of spontaneous mutation and recombination (Aguilera and Klein 1988; Gordenin et al. 1992; Ruskin and Fink 1993; Zou and Rothstein 1997; Kirchner et al. 2000; Pavlov et al. 2001; Galli et al. 2003; Fortune et al. 2005), supporting the link between defective DNA replication and genome instability. In particular, mutations in the POL3 gene that confer a temperature-sensitive growth defect, most likely by affecting the capacity of the cell to replicate its DNA, also confer elevated rates of spontaneous mutation and recombination with a variety of assays (Gordenin et al. 1992, 1993; Tranet al. 1995, 1996, 1997, 1999; Kokoska et al. 1998; Schweitzer and Livingston 1999; Kokoska et al. 2000; Jin et al. 2001; Galli et al. 2003). One of these mutations, pol3-t, is thought to affect the processivity of Pol δ (Gordenin et al. 1992; Tran et al. 1995; Kokoska et al. 2000), which is likely to increase the formation of daughter strand gaps that may be intermediates in the formation of spontaneousmutation and recombination events (Horiuchi et al. 1994; Ivessa et al. 2000; Michel et al. 2001; Minesinger and Jinks-Robertson 2005; Saleh-Gohari et al. 2005; Lopes et al. 2006).

Failure of a replicative polymerase due to an encounter with a spontaneous or induced DNA lesion that blocks its progress provokes a variety of error-free and error-prone responses mediated by a combination of Rad18- and Rad5-dependent post-replication repair and Rad51-dependent recombination repair (Liefshitz et al. …

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