MRC Laboratory of Molecular Biology, Cambridge, UK.
Bioanalytics, Institute of Biotechnology, Technische Universität Berlin, Berlin, Germany.
Nat Struct Mol Biol. 2020 Mar;27(3):240-248. doi: 10.1038/s41594-020-0380-1. Epub 2020 Feb 17.
Vertebrate DNA crosslink repair excises toxic replication-blocking DNA crosslinks. Numerous factors involved in crosslink repair have been identified, and mutations in their corresponding genes cause Fanconi anemia (FA). A key step in crosslink repair is monoubiquitination of the FANCD2-FANCI heterodimer, which then recruits nucleases to remove the DNA lesion. Here, we use cryo-EM to determine the structures of recombinant chicken FANCD2 and FANCI complexes. FANCD2-FANCI adopts a closed conformation when the FANCD2 subunit is monoubiquitinated, creating a channel that encloses double-stranded DNA (dsDNA). Ubiquitin is positioned at the interface of FANCD2 and FANCI, where it acts as a covalent molecular pin to trap the complex on DNA. In contrast, isolated FANCD2 is a homodimer that is unable to bind DNA, suggestive of an autoinhibitory mechanism that prevents premature activation. Together, our work suggests that FANCD2-FANCI is a clamp that is locked onto DNA by ubiquitin, with distinct interfaces that may recruit other DNA repair factors.
脊椎动物 DNA 交联修复切除有毒的复制阻断 DNA 交联。已经鉴定出许多参与交联修复的因素，其相应基因的突变会导致范可尼贫血症（FA）。交联修复的关键步骤是 FANCD2-FANCI 异二聚体的单泛素化，然后招募核酸内切酶去除 DNA 损伤。在这里，我们使用 cryo-EM 确定重组鸡 FANCD2 和 FANCI 复合物的结构。当 FANCD2 亚基单泛素化时，FANCD2-FANCI 采用封闭构象，形成一个封闭双链 DNA（dsDNA）的通道。泛素位于 FANCD2 和 FANCI 的界面处，作为一种共价分子销钉将复合物固定在 DNA 上。相比之下，分离的 FANCD2 是一种无法结合 DNA 的同源二聚体，这表明存在一种自动抑制机制，可防止过早激活。总之，我们的工作表明，FANCD2-FANCI 是一种夹钳，通过泛素锁定在 DNA 上，具有可能招募其他 DNA 修复因子的独特界面。
