Spontaneous Mutations Occur Near Dam Recognition Sites in a dam^- Escherichia coli Host

Margaretha Carraway ¹, Philip Youderian ², and M. G. Marinus ¹

¹ Department of Pharmacology, University of Massachusetts Medical School, Worcester, Massachusetts 01605
² Department of Biological Sciences, University of Southern California, Los Angeles, California 90089

The mismatch repair system of Escherichia coli K12 removes mispaired bases from DNA. Mismatch repair can occur on either strand of DNA if it lacks N⁶-methyladenines within 5'-GATC-3' sequences. In hemimethylated heteroduplexes, repair occurs preferentially on the unmethylated strand. If both strands are fully methylated, repair is inhibited. Mutant (dam^-) strains of E. coli defective in the adenine methylase that recognizes 5'-GATC-3' sequences (Dam), and therefore defective in mismatch repair, show increased spontaneous mutation rates compared to otherwise isogenic dam⁺ hosts. We have isolated and characterized 91 independent mutations that arise as a consequence of the Dam^- defect in a plasmid-borne phage P22 repressor gene, mnt. The majority of these mutations are A:TG:C transitions that occur within six base pairs of the two 5'-GATC-3' sequences in the mnt gene. In contrast, the spectrum of mnt^- mutations in a dam⁺ host is comprised of a majority of insertions of IS elements and deletions that do not cluster near Dam recognition sites. These results show that Dam-directed post-replicative mismatch repair plays a significant role in the rectification of potential transition mutations in vivo, and suggest that sequences associated with Dam recognition sites are particularly prone to replication or repair errors.