REV3 is required for spontaneous but not methylation damage-induced mutagenesis of Saccharomyces cerevisiae cells lacking O-6-methylguanine DNA methyltransferase

O-6-methylguanine (O-6-MeG) DNA methyltransferase (MTase) removes the methy l 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 agent s, but also exhibit an increased spontaneous mutation rate. In order to und erstand molecular mechanisms of endogenous DNA alkylation damage and its ef fects on mutagenesis, we determined the spontaneous mutational spectra of I :he SUP4-o gene in various Saccharomyces cerevisiae strains. To our surpris e, the mgt1 mutant deficient in DNA repair MTase activity exhibited a signi ficant increase in G:C --> C:G transversions instead of the expected G:C -- > A:T transition. Its mutational distribution strongly resembles that of th e rad52 mutant defective in DNA recombinational repair. The rad52 mutationa l spectrum has been shown to be dependent on a mutagenesis pathway mediated by REV3. We demonstrate here that the mgt1 mutational spectrum is also REV 3-dependent and that the rev3 deletion offsets the increase of the spontane ous 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 rev 3 deletion does not affect methylation damage-induced killing and mutagenes is of the mgt1 mutant, suggesting that endogenous alkyl lesions may be diff erent from O-6-MeG. (C) 1999 Elsevier Science B.V. All rights reserved.