Roles for Caenorhabditis elegans rad-51 in Meiosis and in Resistance to Ionizing Radiation During Development

Abstract

We have investigated the role of Caenorhabditis elegans RAD-51 during meiotic prophase and embryogenesis, making use of the silencing effect of RNA interference (RNAi). rad-51 RNAi leads to severe defects in chromosome morphology in diakinesis oocytes. We have explored the effect of rad-51 RNAi in mutants lacking fundamental components of the recombination machinery. If double-strand breaks are prevented by spo-11 mutation, rad-51 RNAi does not affect chromosome appearance. This is consistent with a role for RAD-51 downstream of the initiation of recombination. In the absence of MRE-11, as in the absence of SPO-11, RAD-51 depletion has no effect on the chromosomes, which appear intact, thus indicating a role for MRE-11 in DSB induction. Intriguingly, rad-51 silencing in oocytes that lack MSH-5 leads to chromosome fragmentation, a novel trait that is distinct from that seen in msh-5 mutants and in rad-51 RNAi oocytes, suggesting new potential roles for the msh-5 gene. Silencing of the rad-51 gene also causes a reduction in fecundity, which is suppressed by mutation in the DNA damage checkpoint gene rad-5, but not in the cell death effector gene ced-3. Finally, RAD-51 depletion is also seen to affect the soma, resulting in hypersensitivity to ionizing radiation in late embryogenesis.

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

“…Unlike the brc-1;him-14/MSH4 double mutant, approximately 25% of the observed brc-1;syp-2 nuclei at diakinesis resolve in misshapen, poorly condensed chromatin (Fig 1). This phenotype is similar to that observed in brc-2-and rad-51-deficient strains, which are compromised for repair of all meiotic DSBs through the homologous recombination pathway (Rinaldo et al, 2002). These data indicate that in the absence of crossing over, BRC-1 contributes to meiotic DSB repair by using the sister chromatid as a template.…”

Self-DNA Exposure Induces Developmental Defects and Germline DNA Damage Response in Caenorhabditis elegans

“…In the “mitotic” zone, located at the distal end of each gonadal arm, reside proliferating germ cells, which enter meiosis in the following region, the “transition zone”, containing the leptotene/zygotene stages of meiotic prophase I. Meiotic DNA recombination is initiated in this region upon the formation of DSBs by the conserved topoisomerase subunit SPO-11 [ 28 ]. These DSBs are resected to generate 3′ ssDNA overhangs that are recruited by RAD-51, the eukaryotic RecA homologue, which catalyses the strand-invasion step to mediate repair by interhomolog recombination [ 29 ]. The “transition zone” is followed by the “pachytene” region, where the synaptonemal complex is completely formed and homologous chromosomes are aligned [ 30 , 31 ].…”

“…Lack of evidence of meiotic progression alteration in the presence of activation of the DNA damage checkpoint (above) prompted us to analyse the impact of the self library in spo-11 mutant worms. In the absence of SPO-11, meiotic DSBs are not formed; consequently, RAD-51 is not recruited to chromosomes, and diakinesis nuclei show 12 DAPI-stained bodies corresponding to twelve univalents, in place of six bivalents consisting of homolog chromosomes held together by chiasmata [ 28 , 29 ] ( Figure 6 a). Strikingly, in spo-11 mutant worms fed on the self library, approximatively 16% of oocyte nuclei showed eleven DAPI-stained bodies, corresponding to ten univalents and one bivalent, and 5% of oocyte nuclei showed ten DAPI-stained bodies, corresponding to eight univalents and two bivalents ( Figure 6 a,b).…”

Differential requirement for BRCA1-BARD1 E3 ubiquitin ligase activity in DNA damage repair and meiosis in the Caenorhabditis elegans germ line

“…In addition to enhancing embryonic viability, BRC-1 and BRD-1 stabilize the RAD-51 filament at mid to late pachytene in the zim-1 mutant [41,42]. The RAD-51 recombinase assembles on resected single strand DNA at DSBs and is essential for homology search and strand invasion during homologous recombination [51][52][53]. In wild type, RAD-51 filaments, visualized as nuclear foci by immunostaining, appear in the transition zone (leptotene/zygotene; zone 1), peak in early (zone 2) to mid pachytene (zone 3) and are removed in late pachytene (zones 3-4) [53] (Fig 2B , top).…”