Molecular Mechanism Of TIP60-Mediated XPF-ERCC1 Complex Assembly And Activation

When human genomic DNA is exposed to ultraviolet(UV)radiation or cancer chemotherapeutic agents such as mitomycin C(MMC),covalent links between bases of the same DNA strand(Intrastrand crosslinks)or of the opposite DNA strands(Interstrand crosslinks,ICLs)can be produced.These DNA lesions directly block replication and transcription,thereby resulting in genome instability,eventually leading to xeroderma pigmentosum,Fanconi anemia and other diseases.To cope with the detrimental effects of DNA damage,living organisms have evolved a range of highly conserved surveillance and repair mechanisms.Intrastrand crosslinks are corrected by nucleotide excision repair(NER),while ICLs are repaired by interstrand crosslink(ICL)repair.The XPF-ERCC1 heterodimer is a structure-specific endonuclease that is essential for NER and ICL repair in mammalian cells.Mutations in XPF-ERCC1 have been associated with several human inherited disorders,including Cockayne syndrome,xeroderma pigmentosum,cerebro-oculo-facio-skeletal syndrome,and Fanconi anemia.Biochemical and structural studies have revealed that XPF and ERCC1 form a heterodimer in eukaryotic cells through the interaction of their conserved C-terminal tandem helix-hairpin-helix(Hh H)motifs and the two subunits are unstable without each other.The catalytic subunit XPF has complete nuclease activity only when it forms a tight complex with ERCC1.However,whether and how XPF binding to ERCC1 is regulated has not yet been established.TIP60 is an acetyltransferase involved in the regulation of a wide variety of cellular activities,including gene transcription,chromatin remodeling,and DNA damage repair.TIP60 has also been suggested to regulate the cellular response to UV irradiation,for instance,TIP60 is autoacetylated in response to UV irradiation,which is critically important for TIP60 activation;UV irradiation strongly induced TIP60 expression and stabilization;SUMOylation of TIP60 promotes its acetyltransferase activity in response to UV irradiation.However,the exact role of its acetyltransferase activity in UV damage response remains largely unexplored.Moreover,another study showed that TIP60 directly interacted with the Fanconi anemia protein FANCD2 and was critical for ICL repair.However,downregulation of TIP60 did not affect FANCD2mono-ubiquitination or its localization to DNA damage sites following MMC treatment,indicating that TIP60 may facilitate ICL repair by a yet-unknown mechanism.In this study,we show that TIP60 directly acetylates XPF at Lys911 following UV irradiation or MMC treatment and that this acetylation is required for XPF-ERCC1 complex assembly and subsequent activation.Mechanistically,acetylation of XPF at Lys911 disrupts the Glu907-Lys911 salt bridge,thereby leading to exposure of a previously unidentified second binding site for ERCC1.Accordingly,loss of XPF acetylation impairs the damage-induced XPF-ERCC1 interaction,resulting in defects in both NER and ICL repair.Our results not only reveal a novel mechanism that regulates XPF-ERCC1 complex assembly and activation,but also provide important mechanistic insight into the role of TIP60 in the maintenance of genome stability.