Live-cell single-molecule tracking highlights requirements for stable Smc5/6 chromatin association in vivo.
Live-cell single-molecule tracking highlights requirements for stable Smc5/6 chromatin association in vivo.
复制标题
活细胞单分子追踪强调了体内稳定Smc 5/6染色质缔合的要求。
DOI：
10.7554/elife.68579
复制
发表时间：
2021-04-16
期刊：
影响因子：
7.7
通讯作者：
Murray JM
中科院分区：
文献类型：
--
作者：
Etheridge TJ;Villahermosa D;Campillo-Funollet E;Herbert AD;Irmisch A;Watson AT;Dang HQ;Osborne MA;Oliver AW;Carr AM;Murray JM
必需的Smc 5/6复合物是响应复制应激所必需的，并且最为人所知的是确保同源重组的保真度。使用单分子跟踪活裂酵母调查Smc 5/6染色质协会，我们表明，Smc 5/6是染色质相关的未受挑战的细胞，这取决于非SMC蛋白Nse 6。我们定义了至少两个Nse 6依赖的子途径，其中之一需要BRCT结构域蛋白Brc 1。使用定义的突变体的基因编码的核心Smc 5/6复杂的亚基，我们表明，Nse 3双链DNA结合活性和精氨酸手指的两个Smc 5/6 ATP酶结合位点是至关重要的染色质协会。有趣的是，破坏铰链区的单链DNA（ssDNA）结合活性不会阻止染色质缔合，但会导致复制重新启动期间总染色体重排水平升高。这与ssDNA结合在调节同源重组中的下游功能一致。
The essential Smc5/6 complex is required in response to replication stress and is best known for ensuring the fidelity of homologous recombination. Using single-molecule tracking in live fission yeast to investigate Smc5/6 chromatin association, we show that Smc5/6 is chromatin associated in unchallenged cells and this depends on the non-SMC protein Nse6. We define a minimum of two Nse6-dependent sub-pathways, one of which requires the BRCT-domain protein Brc1. Using defined mutants in genes encoding the core Smc5/6 complex subunits, we show that the Nse3 double-stranded DNA binding activity and the arginine fingers of the two Smc5/6 ATPase binding sites are critical for chromatin association. Interestingly, disrupting the single-stranded DNA (ssDNA) binding activity at the hinge region does not prevent chromatin association but leads to elevated levels of gross chromosomal rearrangements during replication restart. This is consistent with a downstream function for ssDNA binding in regulating homologous recombination.