The structure-specific endonuclease complex SLX4–XPF regulates Tus– Ter -induced homologous recombination

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.