The mutational specificity of two Escherichia coli dnaE antimutator alleles as determined from lacI mutation spectra.
In a companion study we have described the isolation of a series of mutants of Escherichia coli that replicate their DNA with increased fidelity. These mutants carry a mutation in the dnaE gene, encoding the alpha (polymerase) subunit of DNA polymerase III holoenzyme, which is responsible for the faithful replication of the bacterial chromosome. The mutants were detected as suppressors of the high mutability of a mutL strain (defective in postreplicative mismatch correction), in which mutations may be considered to arise predominantly from errors of DNA replication. To investigate the specificity of these antimutator effects, we have analyzed spectra of forward mutations in the N-terminal part of the lacI gene (i-d mutations) for two of the mutL dnaE derivatives (dnaE911 and dnaE915), as well as the control mutL strain. DNA sequencing of over 600 mutants revealed that in the mutL background both antimutator alleles reduce specifically transition mutations (A.T-->G.C and G.C-->A.T). However, the two alleles behave differently in this respect. dnaE911 reduces A.T-->G.C more strongly than it does G.C-->A.T, whereas the reverse is true for dnaE915. Second, dnaE911 does not appear to affect either transversion or frameshift mutations, whereas dnaE915 displays a distinct mutator effect for both. This mutator effect of dnaE915 for frameshift mutations was confirmed by the frequency of reversion of the trpE9777 frameshift mutation. The discovery that dnaE antimutator alleles possess distinct specificities supports the notion that DNA polymerases discriminate against errors along multiple pathways and that these pathways can be influenced independently.
Laboratory of Molecular Genetics, National Institute of Environmental Health Sciences, Research Triangle Park, North Carolina 27709.
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D000483:Alleles; D001426:Bacterial Proteins / Q000235:genetics*; D017122:Bacteriophage T4 / Q000201:enzymology; D001483:Base Sequence; D004258:DNA Polymerase III / Q000235:genetics*; D004269:DNA, Bacterial; D004259:DNA-Directed DNA Polymerase / Q000235:genetics; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics*; D029968:Escherichia coli Proteins; D016368:Frameshift Mutation; D005798:Genes, Bacterial; D007700:Kinetics; D007763:Lac Operon; D056968:Lac Repressors; D008969:Molecular Sequence Data; D009154:Mutation; D012097:Repressor Proteins / Q000235:genetics*
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D029968:Escherichia coli Proteins; D056968:Lac Repressors; C539064:LacI protein, E coli; D012097:Repressor Proteins; C074904:DNA polymerase III, alpha subunit; D004258:DNA Polymerase III; D004259:DNA-Directed DNA Polymerase