REV3 is required for spontaneous but not methylation damage-induced mutagenesis of Saccharomyces cerevisiae cells lacking O6-methylguanine DNA methyltransferase.
O6-methylguanine (O6-MeG) DNA methyltransferase (MTase) removes the methyl group from a DNA lesion and directly restores DNA structure. It has been shown previously that bacterial and yeast cells lacking such MTase activity are not only sensitive to killing and mutagenesis by DNA methylating agents, but also exhibit an increased spontaneous mutation rate. In order to understand molecular mechanisms of endogenous DNA alkylation damage and its effects on mutagenesis, we determined the spontaneous mutational spectra of the SUP4-o gene in various Saccharomyces cerevisiae strains. To our surprise, the mgt1 mutant deficient in DNA repair MTase activity exhibited a significant increase in G:C-->C:G transversions instead of the expected G:C-->A:T transition. Its mutational distribution strongly resembles that of the rad52 mutant defective in DNA recombinational repair. The rad52 mutational spectrum has been shown to be dependent on a mutagenesis pathway mediated by REV3. We demonstrate here that the mgt1 mutational spectrum is also REV3-dependent and that the rev3 deletion offsets the increase of the spontaneous mutation rate seen in the mgt1 strains. These results indicate that the eukaryotic mutagenesis pathway is directly involved in cellular processing of endogenous DNA alkylation damage possibly by the translesion bypass of lesions at the cost of G:C-->C:G transversion mutations. However, the rev3 deletion does not affect methylation damage-induced killing and mutagenesis of the mgt1 mutant, suggesting that endogenous alkyl lesions may be different from O6-MeG.
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Department of Microbiology and Immunology, University of Saskatchewan, Saskatoon, Canada. xiaow@sask.usask.ca
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D001483:Base Sequence; D004249:DNA Damage / Q000235:genetics; D019175:DNA Methylation; D004259:DNA-Directed DNA Polymerase / Q000235:genetics / Q000378:metabolism*; D005656:Fungal Proteins / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D008769:Methylnitronitrosoguanidine / Q000633:toxicity; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009154:Mutation; D019853:O(6)-Methylguanine-DNA Methyltransferase / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins
D005656:Fungal Proteins; D029701:Saccharomyces cerevisiae Proteins; D008769:Methylnitronitrosoguanidine; D019853:O(6)-Methylguanine-DNA Methyltransferase; D004259:DNA-Directed DNA Polymerase; C092602:REV3 protein, S cerevisiae