A MUTATION IN THE MSH5 GENE RESULTS IN ALKYLATION TOLERANCE

DNA methylating agents such as N-methyl-N'-nitro-N-nitrosoguanidine (M NNG) are potent carcinogens; their carcinogenic effect is mainly due t o the effect of production of O-6-methylguanine (O-6 MeG) on DNA. O-6 MeG is not only mutagenic hut also toxic to the cell because Mer(-)/Me x(-) cells unable to remove O-6 MeG are very sensitive to killing by M NNG. It has been proposed that repeated futile mismatch correction of O-6 MeG-containing bp is responsible for the genotoxicity of the O-6 M eG lesion and that loss of mismatch repair activity results in cellula r tolerance to O-6 MeG, but the hypothesis has not been proved. We use d yeast as a model to test this hypothesis and found that chromosome d eletion of any known nuclear mitotic mismatch repair genes, including MLH1, MSH2, MSH3, MSH6, and PMS1, did not rescue mgt1 Delta O-6 MeG DN A repair methyltransferase-deficient cells from killing by MNNG. A lar ge number of mgt1 Delta, MNNG-tolerant revertants were isolated, among which one cell line, XS-14, has been found to carry a mutated allele of the MSH5 gene. The mutation also affected spore survival but did no t increase the spontaneous mutation rate. We further demonstrated that a mutated form of the MSH5 gene, msh5-14, not the msh5 Delta-null mut ation, is responsible for the cellular tolerance to MNNG in XS-14 cell s. This observation offers an alternative model that may reconcile see mingly contradictory observations of yeast and mammalian cells.