Elevated intracellular dCTP levels reduce the induction of GC -> AT transitions in yeast by ethyl methanesulfonate or N-methyl-N'-nitro-N-nitrosoguanidine but increase alkylation-induced GC -> CG transversions

The effect of an increased intracellular dCTP:dTTP ratio onthe specificities of ethyl methanesulfonate (EMS) and N-methyl-N'-nitro-N-nitrosoguanidine(MNNG) mutagenesis was examined in the yeast Saccharomyces cerevisiae.To do so, we used a dCMP deaminase-deficient (ded1) strain havinga dCTP:dTTP ratio >77-fold larger than its isogenic wild-typeparent under the treatment conditions employed. This DNA precursorimbalance lowered the frequencies of EMS- or MNNG-induced SUP4-omutations by 75 or 45% respectively, relative to the correspondingvalues for the wild-type strain. A total of 405 SUP4-o mutationsproduced by the alkylating agents in the dcd1 background werecharacterized by DNA sequencing and the mutational spectra werecompared to those for 399 mutations induced in the wild-typeparent and 207 mutations that arose spontaneously in the dcd1strain. Unexpectedly, the frequencies of EMS- and MNNG-inducedGC AT transitions in the dcd1 strain were found to be reducedby 93 and 68%, respectively, considerably more than the decreasesfor the overall SUP4-o mutation frequencies. The differenceswere due mainly to substantial increases in the frequenciesof GC CG transversions. Although these events were the predominanttype of spontaneous substitution in the dcd1 strain, they weremore frequent after alkylation treatment and were distributeddifferently than the spontaneous GC CG transversions. Preferencesfor the EMS- or MNNG-induced GC AT transitions to occur atGC sites having the guanine located on the transcribed strandor preceded by a 5' purine, respectively, also were diminishedin the dcd1 strain. Together, these findings support mispairingof O⁶-alkylguanine with thymine as the cause of most EMS andMNNG mutagenesis in wild-type yeast. However, the data alsolead us to suggest that the normally error-free repair of apurinicsites resulting from the lability or removal of EMS- or MNNG-inducedN⁷-alkylguanine can be rendered errorprone by increasing thedCTP pool. ¹Present address: Department of Biology, The University of Saskatchewan,Saskatoon, Saskatchewan S7N 0W0, Canada ²To whom correspondence should be addressed