Abstract
uring repair of a double-stranded chromosome break (DSB), an essential step of homologous recombination is searching for homologous DNA sequences and invading the target to repair the break. In somatic (mitotic) cells, this process is performed by the Rad51 recombinase. In meiosis, recombination depends on a meiosis-specific recombinase Dmc1, but still using Rad51 as an accessory factor. Properties of the recombinases have been characterized in vitro, but their cellular strategies working along with relevant mismatch repair proteins remained unexplored in meiosis.
In budding yeast, we compared break-induced replication (BIR) events in both a mitotic and meiotic context, in which a site-specific DSB was created by the HO endonuclease. The invading stand and its donor template share a 108 bp region of homology. We used a series of strains in which the donor carries increasing densities of evenly spaced mismatches. We found that the divergent donor substrates diminished BIR events two times faster in meiosis mediated by Dmc1 comparing to the same events in Rad51-mediated diploid and haploid mitosis. This result differs from in vitro studies that had suggested that Dmc1 had a higher tolerance for mismatches than Rad51. In the absence of Dmc1 and without the meiosis-specific Rad51 inhibitor Hed1, Rad51 was able to carry out ectopic recombination with mismatch tolerance and assimilation similar to the mitotic level.
I also examined the assimilation of mismatches into the BIR repair product. Nearly all mismatch correction in meiosis depends on the proofreading activity of DNA polymerase Polδ and is independent from Msh2. However, homozygous knockout of the mismatch repair protein Msh2 in strains with mismatches elevated the repair rate by nine-fold, but the trend of mismatch correction is the almost same as the wild type. There is no such suppression of recombination of mismatched substrates when Msh2 is deleted in mitotic cells. Collectively, we concluded that both Dmc1 and Msh2 are factors that uniquely shape the recombination patterns in meiosis. Given the high tolerance of Dmc1 validated in vitro is true, we hypothesized that Msh2 might promotes heteroduplex rejection during meiosis to counterbalance the higher mismatch tolerance of Dmc1 when forming the D-loop.