CEA DSV/IRCM, Unité Mixte de Recherche 217 Radiobiologie Moléculaire et Cellulaire, Centre National de la Recherche Scientifique, Commissariat à l'Energie Atomique et aux Energies Alternatives, Fontenay aux Roses, France.
PLoS Genet. 2011 Sep;7(9):e1002305. doi: 10.1371/journal.pgen.1002305. Epub 2011 Sep 29.
Meiotic DNA double-strand breaks (DSBs) initiate crossover (CO) recombination, which is necessary for accurate chromosome segregation, but DSBs may also repair as non-crossovers (NCOs). Multiple recombination pathways with specific intermediates are expected to lead to COs and NCOs. We revisited the mechanisms of meiotic DSB repair and the regulation of CO formation, by conducting a genome-wide analysis of strand-transfer intermediates associated with recombination events. We performed this analysis in a SK1 × S288C Saccharomyces cerevisiae hybrid lacking the mismatch repair (MMR) protein Msh2, to allow efficient detection of heteroduplex DNAs (hDNAs). First, we observed that the anti-recombinogenic activity of MMR is responsible for a 20% drop in CO number, suggesting that in MMR-proficient cells some DSBs are repaired using the sister chromatid as a template when polymorphisms are present. Second, we observed that a large fraction of NCOs were associated with trans-hDNA tracts constrained to a single chromatid. This unexpected finding is compatible with dissolution of double Holliday junctions (dHJs) during repair, and it suggests the existence of a novel control point for CO formation at the level of the dHJ intermediate, in addition to the previously described control point before the dHJ formation step. Finally, we observed that COs are associated with complex hDNA patterns, confirming that the canonical double-strand break repair model is not sufficient to explain the formation of most COs. We propose that multiple factors contribute to the complexity of recombination intermediates. These factors include repair of nicks and double-stranded gaps, template switches between non-sister and sister chromatids, and HJ branch migration. Finally, the good correlation between the strand transfer properties observed in the absence of and in the presence of Msh2 suggests that the intermediates detected in the absence of Msh2 reflect normal intermediates.
减数分裂 DNA 双链断裂 (DSB) 引发交叉 (CO) 重组，这对于准确的染色体分离是必要的，但 DSB 也可能作为非交叉 (NCO) 修复。预计具有特定中间产物的多种重组途径将导致 CO 和 NCO 的形成。我们通过对与重组事件相关的链转移中间产物进行全基因组分析，重新研究了减数分裂 DSB 修复和 CO 形成的调控机制。我们在缺乏错配修复 (MMR) 蛋白 Msh2 的 SK1×S288C 酿酒酵母杂种中进行了这项分析，以允许有效检测异源双链 DNA (hDNA)。首先，我们观察到 MMR 的抗重组活性导致 CO 数量下降 20%，这表明在 MMR 功能正常的细胞中，当存在多态性时，一些 DSB 会使用姐妹染色单体作为模板进行修复。其次，我们观察到很大一部分 NCO 与限制在单个染色单体上的跨 hDNA 片段相关。这一意外发现与修复过程中双链 Holliday 结 (dHJ) 的溶解兼容，并表明除了之前在 dHJ 形成步骤之前描述的控制点之外，在 dHJ 中间产物水平上存在 CO 形成的新控制点。最后，我们观察到 CO 与复杂的 hDNA 模式相关，证实经典的双链断裂修复模型不足以解释大多数 CO 的形成。我们提出，多种因素导致了重组中间产物的复杂性。这些因素包括缺口和双链缺口的修复、非姐妹和姐妹染色单体之间的模板转换，以及 HJ 分支迁移。最后，在缺乏 Msh2 和存在 Msh2 的情况下观察到的链转移特性之间的良好相关性表明，在缺乏 Msh2 的情况下检测到的中间产物反映了正常的中间产物。
