Meiotic crossover control - from DNA breakage to DNA repair

Abstract

Crossovers, important for meiotic homolog segregation, are initiated by DNA double-strand breaks (DSBs) made by Rec12 (Spo11 homolog) aided by a dozen other proteins. Meiosis-specific sister chromatid cohesin subunits Rec8 and Rec11 are loaded nearly uniformly across the genome and promote the binding of linear element proteins Rec10 uniformly with some enrichment at DSB hotspots and Rec25-Rec27-Mug20 with exceptionally high enrichment at hotspots. Elimination of Rec25-Rec27-Mug20 eliminates nearly all DSBs at most hotspots. Thus, these three proteins are determinants of and required for genome-wide DSB hotspots, the first such proteins reported in any species. DSBs are not made independently but interfere with each other over ~100 kb. We propose that hotspot sites are spatially clustered and exactly one DSB is made in each cluster. We find by a type of chromosome conformation capture (“Hi-C”) analysis that DNA from a Rec27-immunoprecipitate ligates preferentially to genomic DNA nearby (<100 kb) and dependent on Rec8, as predicted. Rec12 attaches covalently to DSB ends and is clipped off by the endonuclease of the Mre11-Rad50-Nbs1(MRN)-Ctp1 complex. “Deep sequencing” of the resulting Rec12-oligo complexes reveals the distribution of all DSBs at nearly single-nucleotide resolution. Because there is very low background, this method provides the near absolute level of DSBs across the genome, including low-level DSBs in cold regions, which collectively are abundant. We proposed that crossover invariance, nearly constant cM/kb in spite of DSB hotspots, results from partner choice for DSB repair – primarily with the sister at hotspots and with the homolog in DSB-cold regions. Our analysis of Rec12-oligos supports this interpretation across the genome.