Abstract
In many eukaryotes, including both budding yeast and mammals, repair of double-strand breaks (DSBs) is carried out by different apparatus in somatic and meiotic cells. In mitotic cells, Rad51 recombinase, acting with Rad54, facilitates the search for homology and DNA strand exchange. In meiosis, Rad51 is inhibited by Hed1 and plays only an effector role, while strand exchange is driven by Rad51’s homolog, Dmc1, acting with Rad54’s homolog, Rdh54/Tid1. To directly compare the activities of Rad51 and Dmc1 and especially their tolerance for recombination between divergent sequences, we created diploids in which a site-specific DSB was created by HO endonuclease, either under control of a galactose-inducible promoter or a meiosis-specific SPO13 promoter. Homologous recombination was measured by an ectopic break-induced replication (BIR) assay in which a 108-bp homologous sequence shared between the DSB end and the donor sequence could be easily modified. As previously shown for a haploid mitotic strain, BIR efficiency decreased with increasing divergence between donor and recipient, but repair occurred even when every 6th base pair was mismatched. There was little difference in the tolerance of mismatches in mitotic haploids or meiotic diploids; however, there were notable differences in meiotic diploids when recombination was facilitated by Dmc1 or when Rad51 took over from Dmc1 in both hed1Δ and dmc1Δ hed1Δ mutants. We found that Dmc1 and Rad51 are similarly tolerant of mismatches during meiotic recombination in budding yeast. Surveillance of mismatches by the Msh2 mismatch repair protein proved to be Dmc1-specific. In all cases, assimilation of mismatches into the BIR product was dependent on the 3’ to 5’ exonuclease activity of DNA polymerase δ.