日本东京大学Hitoshi Kurumizaka研究团队解析出组装在含DSB位点核小体上的RAD51冷冻电镜结构。该研究于2024年3月20日在线发表于国际一流学术期刊《自然》。

研究人员报告了人类RAD51核小体复合物的冷冻电镜结构，其中RAD51形成环状和丝状构象。在环状形态中，RAD51原体的N端叶域（NLD）排列在RAD51环的外侧，并直接与核小体DNA结合。L1环和L2环面向RAD51环中心孔的活性中心，从而识别包含双链DNA断裂（DSB）位点的核小体连接DNA。在丝状形式中，核小体DNA被RAD51丝状延伸部分剥离，核小体近端RAD51原基的NLD与剩余的核小体DNA和组蛋白结合。

影响RAD51 NLD的核小体结合残基的突变会减少核小体结合，但几乎不影响体外DNA结合。同样，具有相应突变的酵母Rad51突变体在体内的DNA修复中也存在严重缺陷。这些结果揭示了RAD51 NLD的一种意想不到的功能，并解释了RAD51与核小体结合、识别DSB并在染色质中形成活性丝的机制。

据了解，RAD51是真核生物减数分裂重组和有丝分裂修复DSB所需的核心重组酶。然而，RAD51在染色质中的DSB位点上发挥作用的机制一直难以捉摸。

附：英文原文

Title: Cryo-EM structures of RAD51 assembled on nucleosomes containing a DSB site

Author: Shioi, Takuro, Hatazawa, Suguru, Oya, Eriko, Hosoya, Noriko, Kobayashi, Wataru, Ogasawara, Mitsuo, Kobayashi, Takehiko, Takizawa, Yoshimasa, Kurumizaka, Hitoshi

Issue&Volume: 2024-03-20

Abstract: RAD51 is the central eukaryotic recombinase required for meiotic recombination and mitotic repair of double-strand DNA breaks (DSBs)1,2. However, the mechanism by which RAD51 functions at DSB sites in chromatin has remained elusive. Here we report the cryo-electron microscopy structures of human RAD51–nucleosome complexes, in which RAD51 forms ring and filament conformations. In the ring forms, the N-terminal lobe domains (NLDs) of RAD51 protomers are aligned on the outside of the RAD51 ring, and directly bind to the nucleosomal DNA. The nucleosomal linker DNA that contains the DSB site is recognized by the L1 and L2 loops—active centres that face the central hole of the RAD51 ring. In the filament form, the nucleosomal DNA is peeled by the RAD51 filament extension, and the NLDs of RAD51 protomers proximal to the nucleosome bind to the remaining nucleosomal DNA and histones. Mutations that affect nucleosome-binding residues of the RAD51 NLD decrease nucleosome binding, but barely affect DNA binding in vitro. Consistently, yeast Rad51 mutants with the corresponding mutations are substantially defective in DNA repair in vivo. These results reveal an unexpected function of the RAD51 NLD, and explain the mechanism by which RAD51 associates with nucleosomes, recognizes DSBs and forms the active filament in chromatin.

DOI: 10.1038/s41586-024-07196-4

Source: https://www.nature.com/articles/s41586-024-07196-4