The bacteriophage T4 rnh gene encodes T4 RNase H, a relative of a family of flap endonucleases. T4 rnh null mutations reduce burst sizes, increase sensitivity to DNA damage, and increase the frequency of acriflavin resistance (Acr) mutations. Because mutations in the related Saccharomyces cerevisiae RAD27 gene display a remarkable duplication mutator phenotype, we further explored the impact of rnh mutations upon the mutation process. We observed that most Acrmutants in an rnh + strain

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... mutations, whereas only roughly half of the Acr mutants detected in an rnhΔ strain bearac mutations. In contrast to the mutational specificity displayed by most mutators, the DNA alterations of acmutations arising in rnhΔ andrnh + backgrounds are indistinguishable. Thus, the increase in Acr mutants in an rnhΔ background is probably not due to a mutator effect. This conclusion is supported by the lack of increase in the frequency of rImutations in an rnhΔ background. In a screen that detects mutations at both the rI locus and the much largerrII locus, the r frequency was severalfold lower in anrnhΔ background. This decrease was due to the phenotype of rnh rII double mutants, which display an r+plaque morphology but retain the characteristic inability ofrII mutants to grow on λ lysogens. Finally, we summarize those aspects of T4 forward-mutation systems which are relevant to optimal choices for investigating quantitative and qualitative aspects of the mutation process.