We demonstrated recently that dominant negative mutants of rat DNA polymerase  (Pol ) interfere with repair of alkylation damage in Saccharomyces cerevisiae. To identify the alkylation repair pathway that is disrupted by the Pol  dominant negative mutants, we studied the epistatic relationship of the dominant negative Pol  mutants to genes known to be involved in repair of DNA alkylation damage in S. cerevisiae. We demonstrate that the rat Pol  mutants interfere with the base excision repair pathway in S. cerevisiae. In addition, expression of one of the Pol  dominant negative mutants, Pol -14, increases the spontaneous mutation rate of S. cerevisiae whereas expression of another Pol  dominant negative mutant, Pol -TR, does not. Expression of the Pol -14 mutant in cells lacking APN1 activity does not result in an increase in the spontaneous mutation rate. These results suggest that gaps are required for mutagenesis to occur in the presence of Pol -14 but that it is not merely the presence of a gap that results in mutagenesis. Our results suggest that mutagenesis can occur during the gap-filling step of base excision repair in vivo.