DNA Double Strand Break Repair Laboratory, University of Sussex, Brighton BN1 9RQ, UK, Department of Biochemistry and Molecular Biology, Southern Alberta Cancer Research Institute, University of Calgary, Alberta T2N 4N1, Canada, Radiation Biology and DNA Repair Laboratory, Darmstadt University of Technology, 64287 Darmstadt, Germany and Unidad de Investigación, Hospital Universitario de Canarias, Instituto de Tecnologías Biomédicas, Ofra s/n, 38320 La Laguna, Tenerife, Spain.
Nucleic Acids Res. 2013 Nov;41(21):9719-31. doi: 10.1093/nar/gkt729. Epub 2013 Aug 22.
Although DNA non-homologous end-joining repairs most DNA double-strand breaks (DSBs) in G2 phase, late repairing DSBs undergo resection and repair by homologous recombination (HR). Based on parallels to the situation in G1 cells, previous work has suggested that DSBs that undergo repair by HR predominantly localize to regions of heterochromatin (HC). By using H3K9me3 and H4K20me3 to identify HC regions, we substantiate and extend previous evidence, suggesting that HC-DSBs undergo repair by HR. Next, we examine roles for 53BP1 and BRCA1 in this process. Previous studies have shown that 53BP1 is pro-non-homologous end-joining and anti-HR. Surprisingly, we demonstrate that in G2 phase, 53BP1 is required for HR at HC-DSBs with its role being to promote phosphorylated KAP-1 foci formation. BRCA1, in contrast, is dispensable for pKAP-1 foci formation but relieves the barrier caused by 53BP1. As 53BP1 is retained at irradiation-induced foci during HR, we propose that BRCA1 promotes displacement but retention of 53BP1 to allow resection and any necessary HC modifications to complete HR. In contrast to this role for 53BP1 in HR in G2 phase, we show that it is dispensable for HR in S phase, where HC regions are likely relaxed during replication.
尽管 DNA 非同源末端连接修复了 G2 期大多数的 DNA 双链断裂（DSB），但晚期修复的 DSB 会通过同源重组（HR）进行切除和修复。基于与 G1 细胞情况的相似性，之前的工作表明，主要通过 HR 修复的 DSB 主要定位于异染色质（HC）区域。通过使用 H3K9me3 和 H4K20me3 来鉴定 HC 区域，我们证实并扩展了之前的证据，表明 HC-DSB 通过 HR 进行修复。接下来，我们检查了 53BP1 和 BRCA1 在这个过程中的作用。之前的研究表明，53BP1 促进非同源末端连接，抑制 HR。令人惊讶的是，我们证明在 G2 期，53BP1 是 HR 在 HC-DSB 所必需的，其作用是促进磷酸化 KAP-1 焦点的形成。相比之下，BRCA1 对于 pKAP-1 焦点的形成是可有可无的，但可以缓解 53BP1 造成的障碍。由于 53BP1 在 HR 过程中保留在辐射诱导的焦点中，我们提出 BRCA1 促进 53BP1 的置换和保留，以允许切除和任何必要的 HC 修饰来完成 HR。与 53BP1 在 G2 期 HR 中的这种作用相反，我们表明它在 S 期 HR 中是可有可无的，在复制期间 HC 区域可能会放松。
