The C. elegans DSB-2 Protein Reveals a Regulatory Network that Controls Competence for Meiotic DSB Formation and Promotes Crossover Assurance

Simona Rosu,Abby F Dernburg,Karl A Zawadzki,Anne M Villeneuve,Angela L Reese,Diana E Libuda,Ericca L Stamper,Jeff Sekelsky

doi:10.1371/journal.pgen.1003674

Abstract

For most organisms, chromosome segregation during meiosis relies on deliberate induction of DNA double-strand breaks (DSBs) and repair of a subset of these DSBs as inter-homolog crossovers (COs). However, timing and levels of DSB formation must be tightly controlled to avoid jeopardizing genome integrity. Here we identify the DSB-2 protein, which is required for efficient DSB formation during C. elegans meiosis but is dispensable for later steps of meiotic recombination. DSB-2 localizes to chromatin during the time of DSB formation, and its disappearance coincides with a decline in RAD-51 foci marking early recombination intermediates and precedes appearance of COSA-1 foci marking CO-designated sites. These and other data suggest that DSB-2 and its paralog DSB-1 promote competence for DSB formation. Further, immunofluorescence analyses of wild-type gonads and various meiotic mutants reveal that association of DSB-2 with chromatin is coordinated with multiple distinct aspects of the meiotic program, including the phosphorylation state of nuclear envelope protein SUN-1 and dependence on RAD-50 to load the RAD-51 recombinase at DSB sites. Moreover, association of DSB-2 with chromatin is prolonged in mutants impaired for either DSB formation or formation of downstream CO intermediates. These and other data suggest that association of DSB-2 with chromatin is an indicator of competence for DSB formation, and that cells respond to a deficit of CO-competent recombination intermediates by prolonging the DSB-competent state. In the context of this model, we propose that formation of sufficient CO-competent intermediates engages a negative feedback response that leads to cessation of DSB formation as part of a major coordinated transition in meiotic prophase progression. The proposed negative feedback regulation of DSB formation simultaneously (1) ensures that sufficient DSBs are made to guarantee CO formation and (2) prevents excessive DSB levels that could have deleterious effects.

Highlights

For most diploid organisms, the formation of haploid gametes relies on crossover (CO) recombination between homologous chromosomes for accurate chromosome segregation
We identify C. elegans double-strand breaks (DSBs)-2 as a key regulator of DSB formation, and we propose that its association with chromatin is an indicator of DSB competence
Our data are consistent with a model in which meiotic DSB formation is governed by a negative feedback network wherein cells detect the presence of downstream crossover intermediates and respond by shutting down DSB formation, thereby ensuring that sufficient DSBs are made to guarantee crossovers while simultaneously minimizing the threat to genomic integrity

Summary

Introduction

The formation of haploid gametes relies on crossover (CO) recombination between homologous chromosomes for accurate chromosome segregation. By the last stage of meiotic prophase (diakinesis), the SC has disassembled, and chromosomes have further condensed and reorganized to reveal CO-dependent structures called chiasmata, which connect homologous chromosomes and allow them to orient and segregate to opposite poles at the meiosis I division [2]. DSB formation must be tightly regulated to ensure successful meiosis: cells must both turn on DSB formation to achieve interhomolog COs, and turn off DSB formation to allow repair and subsequent chromosome re-organization in preparation for the meiotic divisions. Cells must make enough DSBs to guarantee one CO per chromosome pair, but too many DSBs could lead to unrepaired DNA damage and compromise genomic integrity

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Discussion

Conclusion