BRC-1 acts in the inter-sister pathway of meiotic double-strand break repair

Abstract

The breast and ovarian cancer susceptibility protein BRCA1 is evolutionarily conserved and functions in DNA double-strand break (DSB) repair through homologous recombination, but its role in meiosis is poorly understood. By using genetic analysis, we investigated the role of the Caenorhabditis elegans BRCA1 orthologue (brc-1) during meiotic prophase. The null mutant in the brc-1 gene is viable, fertile and shows the wild-type complement of six bivalents in most diakinetic nuclei, which is indicative of successful crossover recombination. However, brc-1 mutants show an abnormal increase in apoptosis and RAD-51 foci at pachytene that are abolished by loss of spo-11 function, suggesting a defect in meiosis rather than during premeiotic DNA replication. In genetic backgrounds in which chiasma formation is abrogated, such as him-14/MSH4 and syp-2, loss of brc-1 leads to chromosome fragmentation suggesting that brc-1 is dispensable for crossing over but essential for DSB repair through inter-sister recombination.

Figure 1

Increased number of DAPI-stained structures at diakinesis in brc-1;him-14/MSH4 and in brc-1;syp-2 double mutants. Representative images of DAPI-stained oocyte nuclei at diakinesis in the indicated genotypes double mutants. The number in parenthesis within each image indicates the number of DAPI-stained bodies that were detectable through the z stack of the nucleus. Histograms represent quantification of the DAPI-stained bodies. The number (n) of observed nuclei is indicated next to each genotype. The y axis represents the percentage of nuclei in each class and the x axis indicates the number of DAPI-stained bodies. The difference in DAPI-stained bodies between brc-1;him-14/MSH4 and him-14/MSH4, and between brc-1;syp-2 and syp-2 are statistically significant, (see supplementary Table A online). Scale bar, 2 μm. DAPI, 4,6-diamidino-2-phenylindole; Unst, nuclei with unstructured chromatin; wt, wild type.

Figure 2

brc-1 mutants show spo-11-dependent elevation of RAD-51 foci. Representative image of a wild-type germline immunostained with anti-RAD-51 (red) and DNA counterstained with DAPI (blue). Histograms represent quantification of RAD-51 foci in germlines of animals of the indicated genotypes. The y axis represents the percentage of nuclei with the indicated number of foci. The x axis represents the position (zone) along the germline. Statistical analysis was carried out using the Student's t-test. DAPI, 4,6-diamidino-2-phenylindole; wt, wild type.