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Specificity of the mutator caused by deletion of the yeast structural gene (APN1) for the major apurinic endonuclease

Kunz, B.A., Henson, E.S., Roche, H., Ramotar, D., Nunoshiba, T., Demple, B.
Proceedings of the National Academy of Sciences of the United States of America 1994 v.91 no.17 pp. 8165-8169
DNA repair, DNA, mutagenicity, nucleases, nucleotide sequences, Saccharomyces cerevisiae, structural genes, mutation
The loss of bases from cellular DNA occurs via both spontaneous and mutagen-induced reactions. The resulting apurinic/apyrimidinic (AP) sites 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 residues during replication, the A-rule. Dissimilar results have been obtained by transfecting DNA into eukaryotic cells. It seemed possible that these differences might be due to idiosyncrasies of transfection or aberrant replication of the transfecting DNA. The observation that AP endonuclease-deficient strains of the yeast Saccharomyces cerevisiae have elevated spontaneous mutation rates allowed us to determine the mutational specificity of endogenously generated AP sites in nuclear DNA. With the yeast SUP4-o gene as a mutational target, we found that a deficiency in the major yeast AP endonuclease, Apn1, provoked mainly single base-pair substitution; the rate of transposon Ty insertion was also enhanced. The rate of transversion to a G-C pair was increased 10-fold in Apn1-deficient yeast, including a 59-fold increase in the rate of A-T leads to C-G events. In contrast, the rate of transversion to an A-T pair was increased by only 3-fold. A deficiency in N3-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 here show that AP endonuclease-deficient Escherichia coli exhibit a mutator phenotype consistent with the A-rule.