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

Transformation of both prokaryotes and eukaryotes with singlestranded oligonucleoticles can transfer sequence information from the oligonucleoticle to the chromosome. We have studied this process using oligonucleoticles that correct a -1 frameshift mutation in the LYS2 gene of Saccharomyces cerevisiae. We demonstrate that transformation by oligonucleoticles 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 oligonucleoticles 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 oligonucleoticles 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 Mlhl -independent repair of 7-nt, but not 1-nt, loops. Unexpectedly, we find a strand asymmetry of mismatchrepair function; transformation is blocked more efficiently by MutSa 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 MutSa and MutS beta complexes access and/or repair mismatches that arise in the context of DNA replication.