Meiotic DSB repair by homologous recombination occurs via multiple processes defined by distinct decisions points. One important decision involves partner choice, between recombining with the sister chromatid (the dominant repair partner during mitosis) or with the homolog (the homologous chromosome of different parental origin, the preferred partner during meiosis). Another important decision involves recombination pathway choice, between producing crossovers, where flanking chromosome sequences are exchanged, or noncrossovers. Both decisions appear to be made at early steps in meiotic recombination. Work during the current review period has addressed the following: -------Role of the conserved Sgs1-Top3-Rmi1 helicase-topoisomerase complex. Sgs1-Top3-Rmi1 is the yeast homolog of the mammalian BLM helicase-Top3alpha-BLAP75 complex, implicated in cancer avoidance and recombination control in humans. We previously showed that all three members of the yeast complex are essential for normal recombination partner choice and for population of regulated meiotic crossover and noncrossover recombination pathways. Based on these findings, we hypothesized that Sgs1-Top3-Rmi1 acts as a chaperone for early recombination intermediates. We also identified an Sgs1-independent role for Top3-Rmi1, in resolving strand entanglements that accumulate in recombination intermediates. We have now shown, using a procedure where recombination intermediates are accumulated during wild-type meiosis but resolved in the absence of Sgs1, Rmi1, or both, that Sgs1 activity is not required to fully resolve recombination intermediates, but Top3-Rmi1 is. These findings confirm conclusions based on meiotic studies, and extent their applicability to recombination events that occur during the mitotic cell cycle. Current work is aimed at testing the recombination intermediate chaperone hypothesis by examining parental strand contributions to meiotic recombinants. We find that the vast majority of meiotic recombinants contain mosaic parental strand contributions, consistent with a model in which strand invasion can occur on either the sister chromatid or other parental homolog, and in which frequent partner switching, driven by Sgs1-Top3-Rmi1, leads to mosaic recombinants. -------Impact of chromosome structure on recombination biochemistry. Meiotic chromosomes are organized around a protein axis. A subset of axis components, important for meiotic DSB formation and partner choice, are enriched in some regions of the genome relative to others. To examine their impact on recombination biochemistry independent of their role in recombination initiation, we are studying recombination initiated by VDE, a sequence-specific nuclease whose activity is independent of local axis composition. We have no obtained data indicating that local chromosome structure determines the biochemical mechanism of recombination intermediate resolution. At an axis protein-enriched locus, VDE-initiated crossovers form using meiosis-specific resolution activities, but using mitotic resolvases at an axis protein-depleted locus. Current work is aimed at testing the generality of this conclusion at other loci, by altering axis protein enrichment at a specific locus, and also at examining the impact of chromosome structure on earlier, pre-resolution steps in the recombination process.
Showing the most recent 10 out of 17 publications
