Oligonucleotide transformation of yeast reveals mismatch repair complexes to be differentially active on DNA replication strands.

Published

Journal Article

Transformation of both prokaryotes and eukaryotes with single-stranded oligonucleotides can transfer sequence information from the oligonucleotide to the chromosome. We have studied this process using oligonucleotides that correct a -1 frameshift mutation in the LYS2 gene of Saccharomyces cerevisiae. We demonstrate that transformation by oligonucleotides occurs preferentially on the lagging strand of replication and is strongly inhibited by the mismatch-repair system. These results are consistent with a mechanism in which oligonucleotides anneal to single-stranded regions of DNA at a replication fork and serve as primers for DNA synthesis. Because the mispairs the primers create are efficiently removed by the mismatch-repair system, single-stranded oligonucleotides can be used to probe mismatch-repair function in a chromosomal context. Removal of mispairs created by annealing of the single-stranded oligonucleotides to the chromosomal DNA is as expected, with 7-nt loops being recognized solely by MutS beta and 1-nt loops being recognized by both MutS alpha and MutS beta. We also find evidence for Mlh1-independent repair of 7-nt, but not 1-nt, loops. Unexpectedly, we find a strand asymmetry of mismatch-repair function; transformation is blocked more efficiently by MutS alpha on the lagging strand of replication, whereas MutS beta does not show a significant strand bias. These results suggest an inherent strand-related difference in how the yeast MutS alpha and MutS beta complexes access and/or repair mismatches that arise in the context of DNA replication.

Full Text

Duke Authors

Cited Authors

  • Kow, YW; Bao, G; Reeves, JW; Jinks-Robertson, S; Crouse, GF

Published Date

  • July 3, 2007

Published In

Volume / Issue

  • 104 / 27

Start / End Page

  • 11352 - 11357

PubMed ID

  • 17592146

Pubmed Central ID

  • 17592146

International Standard Serial Number (ISSN)

  • 0027-8424

Digital Object Identifier (DOI)

  • 10.1073/pnas.0704695104

Language

  • eng

Conference Location

  • United States