SPECIFICITY OF THE MUTATOR CAUSED BY DELETION OF THE YEAST STRUCTURALGENE (APN1) FOR THE MAJOR APURINIC ENDONUCLEASE

The loss of bases from cellular DNA occurs via both spontaneous and mu tagen-induced reactions. The resulting apurinic/apyrimidinic (AP) site s are cytotoxic and mutagenic but are counteracted by repair initiated by AP endonucleases. Previously, in vitro and bacterial transfection studies suggested that AP sites often prompt insertion of dAMP residue s during replication, the A-rule. Dissimilar results have been obtaine d by transfecting DNA into eukaryotic cells. It seemed possible that t hese differences might be due to idiosyncrasies of transfection or abe rrant replication of the transfecting DNA. The observation that AP end onuclease-deficient strains of the yeast Saccharomyces cerevisiae have elevated spontaneous mutation rates allowed us to determine the mu- t ational specificity of endogenously generated AP sites in nuclear DNA. With the yeast SUP4-o gene as a mutational target, we found that a de ficiency in the major yeast AP endonuclease, Apn1, provoked mainly sin gle base-pair substitution; the rate of transposon Ty insertion was al so enhanced. The rate of transversion to a G C pair was increased 10-f old in Apn1-deficient yeast, including a 59-fold increase in the rate of A.T --> C.G events. In contrast, the rate of transversion to an A.T pair was increased by only 3-foid. A deficiency in N-3-methyladenine glycosylase offset these substitution rate increases, indicating that they are due primarily to AP sites resulting from glycosylase action. Thus, the A-rule does not seem to apply to the mutagenic processing of endogenous abasic sites in S. cerevisiae. Other results presented her e show that AP endonuclease-deficient Escherichia coli exhibit a mutat or phenotype consistent with the A-rule.