Sequence composition and context effects on the generation and repair of frameshift intermediates in mononucleotide runs in Saccharomyces cerevisiae

DNA. polymerase slippage occurs frequently in tracts of a tandemly repeated nucleotide, and such slippage events can be genetically detected as frames hift mutations. In long mononucleotide runs, most frameshift intermediates are repaired by the postreplicative mismatch repair (MMR) machinery, rather than by the exonucleolytic proofreading activity of DNA polymerase. Althou gh mononucleotide runs are hotspots for polymerase slippage events, it is n ot known whether the composition of a run and the surrounding context affec t the frequency of slippage or the efficiency of MMR. To address these issu es, 10-nucleotide (10N) runs were inserted into the yeast LYSE gene to crea te fl frameshift alleles. Slippage events within these runs were detected a s Lys(+) revertants. 10G or 10C runs were found to be more unstable than 10 A or 10T runs, but neither the frequency of polymerase slippage nor the ove rall efficiency of MMR was greatly influenced by sequence context. Although complete elimination of MMR activity (msh2 mutants) affected all runs simi larly, analyses of reversion rates in msh3 and msh6 mutants revealed distin ct specificities of the yeast Msh2p-Msh3p and Msh2p-Msh6p mismatch binding complexes in the repair of frameshift intermediates in different sequence c ontexts.