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

Genetics. 2000 Oct;156(2):571-8. doi: 10.1093/genetics/156.2.571.

Abstract

DNA polymerase slippage occurs frequently in tracts of a tandemly repeated nucleotide, and such slippage events can be genetically detected as frameshift 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. Although mononucleotide runs are hotspots for polymerase slippage events, it is not known whether the composition of a run and the surrounding context affect the frequency of slippage or the efficiency of MMR. To address these issues, 10-nucleotide (10N) runs were inserted into the yeast LYS2 gene to create +1 frameshift alleles. Slippage events within these runs were detected as Lys(+) revertants. 10G or 10C runs were found to be more unstable than 10A or 10T runs, but neither the frequency of polymerase slippage nor the overall efficiency of MMR was greatly influenced by sequence context. Although complete elimination of MMR activity (msh2 mutants) affected all runs similarly, analyses of reversion rates in msh3 and msh6 mutants revealed distinct specificities of the yeast Msh2p-Msh3p and Msh2p-Msh6p mismatch binding complexes in the repair of frameshift intermediates in different sequence contexts.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Aldehyde Oxidoreductases / genetics*
  • Alleles
  • Base Pair Mismatch*
  • Base Sequence
  • DNA Repair*
  • DNA-Binding Proteins / genetics
  • DNA-Directed DNA Polymerase / metabolism
  • Frameshift Mutation*
  • Fungal Proteins / genetics
  • L-Aminoadipate-Semialdehyde Dehydrogenase
  • Molecular Sequence Data
  • MutS Homolog 2 Protein
  • Mutagenesis, Site-Directed
  • Mutation
  • Protein Subunits
  • Saccharomyces cerevisiae / genetics*
  • Saccharomyces cerevisiae Proteins*

Substances

  • DNA-Binding Proteins
  • Fungal Proteins
  • MSH6 protein, S cerevisiae
  • Protein Subunits
  • Saccharomyces cerevisiae Proteins
  • Aldehyde Oxidoreductases
  • L-Aminoadipate-Semialdehyde Dehydrogenase
  • DNA-Directed DNA Polymerase
  • MSH2 protein, S cerevisiae
  • MutS Homolog 2 Protein