Vertebrate replication forks arrested at interstrand DNA cross-links (ICLs) engage the Fanconi anemia pathway to incise arrested forks, ‘unhooking’ the ICL and forming a double strand break (DSB) that is repaired by homologous recombination (HR). The FANCP product, SLX4, in complex with the XPF (also known as FANCQ or ERCC4)–ERCC1 endonuclease, mediates ICL unhooking. Whether this mechanism operates at replication fork barriers other than ICLs is unknown. Here, we study the role of mouse SLX4 in HR triggered by a site-specific chromosomal DNA–protein replication fork barrier formed by the Escherichia coli-derived Tus–Ter complex. We show that SLX4–XPF is required for Tus–Ter-induced HR but not for error-free HR induced by a replication-independent DSB. We additionally uncover a role for SLX4–XPF in DSB-induced long-tract gene conversion, an error-prone HR pathway related to break-induced replication. Notably, Slx4 and Xpf mutants that are defective for Tus–Ter-induced HR are hypersensitive to ICLs and also to the DNA–protein cross-linking agents 5-aza-2′-deoxycytidine and zebularine. Collectively, these findings show that SLX4–XPF can process DNA–protein fork barriers for HR and that the Tus–Ter system recapitulates this process.

脊椎动物复制叉在链间 DNA 交联 (ICL) 处停滞，利用范可尼贫血途径切割停滞的复制叉，“解开”ICL 并形成双链断裂 (DSB)，并通过同源重组 (HR) 进行修复。 FANCP产物 SLX4 与 XPF（也称为 FANCQ 或 ERCC4）-ERCC1 核酸内切酶复合，介导 ICL 脱钩。该机制是否在 ICL 以外的复制叉屏障上起作用尚不清楚。在这里，我们研究了小鼠 SLX4 在由大肠杆菌衍生的 Tus- Ter复合物形成的位点特异性染色体 DNA-蛋白质复制叉屏障触发的 HR 中的作用。我们表明，SLX4–XPF 是 Tus– Ter诱导的 HR 所必需的，但对于复制独立的 DSB 诱导的无错误 HR 则不需要。我们还发现了 SLX4-XPF 在 DSB 诱导的长链基因转换中的作用，这是一种与断裂诱导复制相关的容易出错的 HR 途径。值得注意的是，Tus- Ter诱导的 HR 有缺陷的Slx4和Xpf突变体对 ICL 以及 DNA-蛋白质交联剂 5-aza-2'-deoxycytidine 和 zebularine 过敏。总的来说，这些发现表明 SLX4-XPF 可以处理 HR 的 DNA-蛋白质叉屏障，并且 Tus- Ter系统概括了这一过程。