DNA polymerase III proofreading mutants enhance the expansion and deletionof triplet repeat sequences in Escherichia coli

The influence of mutations in the 3' to 5' exonucleolytic proofreading c-su bunit of Escherichia coli DNA polymerase III on the genetic instabilities o f the CGG.CCG and the CTG.CAG repeats that cause human hereditary neurologi cal diseases was investigated. The dnaQ49(ts) and the mutD5 mutations desta bilize the CGG.CCG repeats. The distributions of the deletion products indi cate that slipped structures containing a small number of repeats in the lo op mediate the deletion process. The CTG.CAG repeats were destabilized by t he dnaQ49(ts) mutation by a process mediated by long hairpin loop structure s (greater than or equal to 5 repeats). The mutD5 mutator strain stabilized the (CTG.CAG)(175) tract, which contained two interruptions. Since the mut D5 mutator strain has a saturated mismatch repair system, the stabilization is probably an indirect effect of the nonfunctional mismatch repair system in these strains. Shorter uninterrupted tracts expand readily in the mutD5 strain, presumably due to the greater stability of long CTG.CAG tracts (>1 00 repeats) in this strain. When parallel studies were conducted in minimal medium, where the mutD5 strain is defective in exonucleolytic proofreading but has a functional MMR system, both CTG.CAG and CGG.CCG repeats were des tabilized, showing that the proofreading activity is essential for maintain ing the integrity of TRS tracts. Thus, we conclude that the expansion and d eletion of triplet repeats are enhanced by mutations that reduce the fideli ty of replication.