title: XPF with mutations in its conserved nuclease domain is defective in DNA repair but functions in TRF2-mediated telomere shortening.
abstract: TRF2, a telomere-binding protein, is a crucial player in telomere length maintenance. Overexpression of TRF2 results in telomere shortening in both normal primary fibroblasts and telomerase-positive cancer cells. TRF2 is found to be associated with XPF-ERCC1, a structure-specific endonuclease involved in nucleotide excision repair, crosslink repair and DNA recombination. XPF-ERCC1 is implicated in TRF2-dependent telomere loss in mouse keratinocytes, however, whether XPF-ERCC1 and its nuclease activity are required for TRF2-mediated telomere shortening in human cells is unknown. Here we report that TRF2-induced telomere shortening is abrogated in human cells deficient in XPF, demonstrating that XPF-ERCC1 is required for TRF2-promoted telomere shortening. To further understand the role of XPF in TRF2-dependent telomere shortening, we generated constructs containing either wild type XPF or mutant XPF proteins carrying amino acid substitutions in its conserved nuclease domain. We show that wild type XPF can complement XPF-deficient cells for repair of UV-induced DNA damage whereas the nuclease-inactive XPF proteins fail to do so, indicating that the nuclease activity of XPF is essential for nucleotide excision repair. In contrast, both wild type XPF and nuclease-inactive XPF proteins, when expressed in XPF-deficient cells, are able to rescue TRF2-mediated telomere shortening. Thus, our results suggest that the function of XPF in TRF2-mediated telomere shortening is conserved between mouse and human. Furthermore, our findings reveal an unanticipated nuclease-independent function of XPF in TRF2-mediated telomere shortening.
authors.affiliation: Department of Biology, LSB438, McMaster University, 1280 Main St. West, Hamilton, Ontario, Canada L8S4K1.
authors.forename: Yili
authors.initials: Y
authors.lastname: Wu
authors.forename: Natalie J
authors.initials: NJ
authors.lastname: Zacal
authors.forename: Andrew J
authors.initials: AJ
authors.lastname: Rainbow
authors.forename: Xu-Dong
authors.initials: XD
authors.lastname: Zhu
mesh_terms: D019943:Amino Acid Substitution; D002460:Cell Line; D017124:Conserved Sequence; D004260:DNA Repair / Q000235:genetics* / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D003851:Deoxyribonucleases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D005816:Genetic Complementation Test; D006801:Humans; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D017434:Protein Structure, Tertiary; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016615:Telomere / Q000235:genetics / Q000378:metabolism*; D035341:Telomeric Repeat Binding Protein 2 / Q000235:genetics / Q000378:metabolism*; D014466:Ultraviolet Rays
chemical_list: D004268:DNA-Binding Proteins; D011994:Recombinant Proteins; D035341:Telomeric Repeat Binding Protein 2; C095888:xeroderma pigmentosum group F protein; D003851:Deoxyribonucleases