The proper segregation of chromosomes during meiosis relies on a series of molecular events leading to the formation of crossovers (COs) between homologous chromosomes. The formation of COs initiates with programmed double-strand breaks (DSBs) in the DNA. After DSB induction, breaks are resected and channeled into homologous recombination (HR) repair rather than to other, more error-prone repair pathways including non-homologous end-joining (NHEJ), alternative NHEJ (alt-NHEJ) and single-strand annealing (SSA). While previous works suggest that NHEJ is prevented by early steps of resection, little is known about the mechanisms inhibiting the other pathways. In C. elegans, meiotic HR relies on the recombinase RAD-51 to invade a homologous template for repair. In wild type worms, HR-mediated repair leads to the formation of 6 pairs of CO-attached homologs at the end of prophase I. When rad-51 is mutated, DSBs are not repaired properly and massive chromosomal fusions occur. We discovered that depleting DSB-promoting factors, such as him-5 or dsb-2, abrogates chromosomal fusions in rad-51 mutants, independently from their role in DSB formation. The diakinesis nuclei of rad-51;him-5 and dsb-2;rad-51 contain 12 intact univalent chromosomes indicating repaired DSBs independently from HR. This data leads us to hypothesize that the DSB machinery has a role in DNA-repair pathway choice during meiosis. We thus investigated the role of NHEJ, alt-NHEJ and SSA during oocyte formation in a rad-51;him-5 double-mutant context. These three pathways are compatible with the use as template of the different DNA intermediates which are formed as a result of the processing of DSB ends. First, we observed that in the absence of rad-51 alone, NHEJ and SSA are the two major repair pathways used by the cell, while alt-NHEJ does not contribute significantly to repair. By contrast, in the rad-51;him-5 double mutant, NHEJ and alt-NHEJ are the major repair pathways. Thus, the absence of him-5 switches the preferred repair pathways from SSA to alt-NHEJ. We propose that him-5 promotes HR by favoring extended resection of the DSB ends, leading to the formation of a template for RAD-51-dependent HR or for SSA in a rad-51 mutant. This work provides insights on the regulation of meiotic DSB-repair pathway choice. In addition to its role in DSB formation, him-5 appears to promote HR by preventing alt-NHEJ-mediated repair, revealing that components of the DSB-promoting machinery inhibit alternative repair to promote HR.

Affiliation:
- Magee-Womens Research Institute, University of Pittsburgh, Pittsburgh, PA