RPA activates the XPF‐ERCC1 endonuclease to initiate processing of DNA interstrand crosslinks
AuthorsAbdullah, Ummi B
McGouran, Joanna F
El‐Sagheer, Afaf H
McHugh, Peter J
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AbstractDuring replication‐coupled DNA interstrand crosslink (ICL) repair, the XPF‐ERCC1 endonuclease is required for the incisions that release, or “unhook”, ICLs, but the mechanism of ICL unhooking remains largely unknown. Incisions are triggered when the nascent leading strand of a replication fork strikes the ICL. Here, we report that while purified XPF‐ERCC1 incises simple ICL‐containing model replication fork structures, the presence of a nascent leading strand, modelling the effects of replication arrest, inhibits this activity. Strikingly, the addition of the single‐stranded DNA (ssDNA)‐binding replication protein A (RPA) selectively restores XPF‐ERCC1 endonuclease activity on this structure. The 5′–3′ exonuclease SNM1A can load from the XPF‐ERCC1‐RPA‐induced incisions and digest past the crosslink to quantitatively complete the unhooking reaction. We postulate that these collaborative activities of XPF‐ERCC1, RPA and SNM1A might explain how ICL unhooking is achieved in vivo.
CitationAbdullah UB, McGouran JF, Brolih S, Ptchelkine D, El‐Sagheer AH, et al. (2017) RPA activates the XPF‐ERCC1 endonuclease to initiate processing of DNA interstrand crosslinks. The EMBO Journal: e201796664. Available: http://dx.doi.org/10.15252/embj.201796664.
SponsorsWe thank our laboratory member Sook Y. Lee and Opher Gileadi (Structural Genomics Consortium, Oxford) for the recombinant SNM1A; Fumiko Esashi (Dunn School of Pathology, University of Oxford) and Marc Wold (Iowa State University, USA) for the RPA and RPA70ΔC442, respectively; Hannah Baddock (WIMM), Hazel Aitkenhead and Joseph A. Newman (Structural Genomics Consortium, University of Oxford) for assistance with fluorescence anisotropy. P.J.M. was supported by MRC (Medical Research Council, grant MR/L007665/1). A.H.E.S. and T.B. were supported by BBSRC (Biotechnology and Biological Sciences Research Council, grant BB/J001694/1) BBSRC: sLOLA grant BB/J001694/1 “Extending the boundaries of nucleic acid chemistry” J.F.M. was supported by King Abdullah University of Science and Technology (KAUST) for a grant awarded to B. Nordén. U.B.A. was supported by the Malaysia's King Scholarship (Biasiswa Yang Di‐Pertuan Agong).
JournalThe EMBO Journal