NOVEL ESSENTIAL DNA REPAIR PROTEINS NSE1 AND NSE2 ARE SUBUNITS OF THE FISSION Y

该子项目是利用NIH/NCRR资助的中心赠款提供的资源的许多研究子项目之一。子项目和研究者（PI）可能从另一个NIH来源获得主要资金，因此可以在其他CRISP条目中表示。所列机构为中心机构，不一定为研究者机构。染色体结构维持蛋白（SMC）家族在基因组稳定性中起着重要作用。SMC异二聚体是姐妹染色单体凝聚（Cohesin：Smc 1和Smc 3）、染色质凝聚（Condensin：Smc 2和Smc 4）和DNA修复（Smc 5和Smc 6）所必需的。SMC异二聚体不能单独发挥作用，必须与必需的非SMC亚基结合。为了进一步了解Smc 5 -6复合物的基本作用和DNA修复作用，我们纯化了裂殖酵母Smc 5，并通过质谱鉴定了共沉淀蛋白Nse 1和Nse 2。我们发现，Nse 1和Nse 2在体内与Smc 5相互作用，作为Smc 5 -6复合物的一部分。Nse 1和Nse 2是酵母和人类的重要蛋白质，其功能的丧失导致了与Smc 5 -6失活所观察到的那些惊人相似的末端表型。此外，表达Nse 1和Nse 2的亚纯型等位基因的细胞与Smc 5 -6突变体一样，对DNA损伤高度敏感。上位性分析表明，与Smc 5 -6一样，Nse 1和Nse 2与Rhp 51一起在DNA双链断裂的同源重组修复中起作用。这项研究的结果强烈表明，Nse 1和Nse 2是新的非SMC亚基的裂殖酵母Smc 5 -6 DNA修复复合物。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. The structural maintenance of chromosomes (SMC) family of proteins play essential roles in genomic stability. SMC heterodimers are required for sister-chromatid cohesion (Cohesin: Smc1 & Smc3), chromatin condensation (Condensin: Smc2 & Smc4), and DNA repair (Smc5 & Smc6). The SMC heterodimers do not function alone and must associate with essential non-SMC subunits. To gain further insight into the essential and DNA repair roles of the Smc5-6 complex, we have purified fission yeast Smc5 and identified by mass spectrometry the co-precipitating proteins, Nse1 and Nse2. We show that both Nse1 and Nse2 interact with Smc5 in vivo, as part of the Smc5-6 complex. Nse1 and Nse2 are essential proteins and conserved from yeast to man. Loss of Nse1 and Nse2 function leads to strikingly similar terminal phenotypes to those observed for Smc5-6 inactivation. In addition, cells expressing hypomorphic alleles of Nse1 and Nse2 are, like Smc5-6 mutants, hypersensitive to DNA damage. Epistasis analysis suggests that like Smc5-6, Nse1, and Nse2 function together with Rhp51 in the homologous recombination repair of DNA double strand breaks. The results of this study strongly suggest that Nse1 and Nse2 are novel non-SMC subunits of the fission yeast Smc5-6 DNA repair complex.