Antimutator strains (ie., strains that have lower mutation rates than the parental wild-type strain) are important tools in elucidating the precise mechanisms by which organisms produce mutations. In general, antimutators can be thought of as organisms that possess increased efficiency in certain cellular mutation-prevention systems. The well-defined genetics and molecular biology of the bacterium E. coli permit a systematic search for mutants that are thus affected. For example, by means of specific tests, mutants may be isolated that produce less errors of replication. Subsequent demonstration that these mutants are antimutators for overall spontaneous mutagenesis provides evidence that errors of DNA replication are a source of spontaneous mutations. Analogously, mutants may be isolated with increased efficiency for defined DNA repair capacities, and decreased mutagenesis of such strains establishes defined group of DNA damages as contributors to spontaneous mutation. Using a papillation assay in mismatch-repair-defective strains, we were successful in isolating mutants of E. coli that replicate their DNA with increased accuracy. The strains were found to carry a mutation in the dnaE gene, which encodes the DNA polymerase III that is primarily responsible for the replication of the bacterial chromosome. When tested in an otherwise wild-type background, the dnaE alleles were also capable of reducing the mutation rates in this background by about 2-fold. This suggest that ~50 % of spontaneous mutations in E. coli may result from uncorrected DNA replication errors. We are also investigating the mechanisms by which these antimutator alleles exert their effect. Our results so far indicate, based on both in vivo and in vitro studies, that the antimutator effects do not likely result from increased base selection by the DNA polymerase. Instead, they may result from (i) impaired catalysis leading to increased exonucleolytic proofreading and (ii) polymerase instability leading to increased dissociation from the replication fork. These studies will likely provide insights into the normal fidelity mechanisms as well.
| Schaaper, R M; Dunn, R L (2001) The antimutator phenotype of E. coli mud is only apparent and results from delayed appearance of mutants. Mutat Res 480-481:71-5 |
