Zhao Song, Renthal William, Lee Eva Y-H P
Department of Molecular Medicine/Institute of Biotechnology, University of Texas Health Science Center, San Antonio, TX 78245, USA.
Nucleic Acids Res. 2002 Nov 15;30(22):4815-22. doi: 10.1093/nar/gkf612.
Rad50/Mre11/NBS1 (R/M/N) is a multi-functional protein complex involved in DNA repair, cell cycle checkpoint activation, DNA replication and replication block-induced responses. Ionizing radiation (IR) induces the phosphorylation of NBS1 and nuclear foci formation of the complex. Although it has been suggested that the R/M/N complex is associated with DNA damage sites, we present here biochemical evidence for chromatin association of the complex. We show that the chromatin association of R/M/N is independent of IR and ataxia telangiectasia mutated (ATM). We also demonstrate that optimal chromatin association of the Rad50/Mre11/NBS1 proteins requires both the conserved forkhead-associated (FHA) and breast cancer C-terminus (BRCT) domains of NBS1. Moreover, both these domains of NBS1 are required for its phosphorylation on Ser343 but not on Ser278. Importantly, both the FHA and BRCT domains are essential for IR-induced foci (IRIF) formation of R/M/N and S phase checkpoint activation, but only the BRCT domain is needed for cell survival after IR. These data demonstrate that the FHA and BRCT domains of NBS1 are crucial for the functions of the R/M/N complex.
Rad50/Mre11/NBS1（R/M/N）是一种多功能蛋白复合物，参与DNA修复、细胞周期检查点激活、DNA复制以及复制阻滞诱导的反应。电离辐射（IR）可诱导NBS1磷酸化以及该复合物形成核灶。尽管有人提出R/M/N复合物与DNA损伤位点有关，但我们在此提供该复合物与染色质结合的生化证据。我们表明，R/M/N与染色质的结合不依赖于IR和共济失调毛细血管扩张症突变基因（ATM）。我们还证明，Rad50/Mre11/NBS1蛋白与染色质的最佳结合需要NBS1保守的叉头相关（FHA）结构域和乳腺癌C端（BRCT）结构域。此外，NBS1的这两个结构域对于其Ser343位点的磷酸化是必需的，但对于Ser278位点的磷酸化则不是必需的。重要的是，FHA和BRCT结构域对于R/M/N的IR诱导灶（IRIF）形成和S期检查点激活都是必不可少的，但IR后细胞存活仅需要BRCT结构域。这些数据表明，NBS1的FHA和BRCT结构域对于R/M/N复合物的功能至关重要。
