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Published March 6, 2019 | Accepted Version + Supplemental Material
Journal Article Open

TRAIP is a master regulator of DNA interstrand crosslink repair


Cells often use multiple pathways to repair the same DNA lesion, and the choice of pathway has substantial implications for the fidelity of genome maintenance. DNA interstrand crosslinks covalently link the two strands of DNA, and thereby block replication and transcription; the cytotoxicity of these crosslinks is exploited for chemotherapy. In Xenopus egg extracts, the collision of replication forks with interstrand crosslinks initiates two distinct repair pathways. NEIL3 glycosylase can cleave the crosslink; however, if this fails, Fanconi anaemia proteins incise the phosphodiester backbone that surrounds the interstrand crosslink, generating a double-strand-break intermediate that is repaired by homologous recombination. It is not known how the simpler NEIL3 pathway is prioritized over the Fanconi anaemia pathway, which can cause genomic rearrangements. Here we show that the E3 ubiquitin ligase TRAIP is required for both pathways. When two replisomes converge at an interstrand crosslink, TRAIP ubiquitylates the replicative DNA helicase CMG (the complex of CDC45, MCM2–7 and GINS). Short ubiquitin chains recruit NEIL3 through direct binding, whereas longer chains are required for the unloading of CMG by the p97 ATPase, which enables the Fanconi anaemia pathway. Thus, TRAIP controls the choice between the two known pathways of replication-coupled interstrand-crosslink repair. These results, together with our other recent findings establish TRAIP as a master regulator of CMG unloading and the response of the replisome to obstacles.

Additional Information

© 2019 Springer Nature Limited. Received 05 July 2018. Accepted 01 February 2019. Published 06 March 2019. We thank D. Pellman and members of the Walter laboratory for comments on the manuscript, and K. Arnett for help with biolayer interferometry experiments. J.C.W. is supported by NIH grant HL098316 and a gift from the family of Jonathan G. Wiseman. R.A.W. is supported by American Cancer Society postdoctoral fellowship 131415-PF-17-168-01-DMC, D.R.S. by NIH award K99GM129422, D.R.S. and G.C. by Jane Coffin Childs postdoctoral fellowships, J.L.S. by a Damon Runyon postdoctoral fellowship, M.W. by the Cancer Research UK Clinician Scientist Fellowship, and E.L. by NIH award F31GM122277. J.C.W. is a Howard Hughes Medical Institute Investigator and an American Cancer Society Research Professor. These authors contributed equally: R. Alex Wu, Daniel R. Semlow Autjhor Contributions. R.A.W. identified TRAIP as the E3 ligase that ubiquitylates CMG and characterized the role of TRAIP in cisplatin-ICL repair. D.R.S. characterized the role of CMG ubiquitylation in AP-ICL repair and performed the structure–function analysis of NEIL3. A.N.K.-L., M.R.H. and M.W. generated Fig. 4d and Extended Data Fig. 9 under the supervision of K.J.P. O.V.K. generated Fig. 1c, d and Extended Data Figs. 2i and 6a, b. G.C. and E.L. prepared Xenopus laevis rCMG. R.A. generated Fig. 1e and Extended Data Fig. 3c, d. J.L.S. generated Fig. 2e and Extended Data Fig. 4c. L.D. generated Extended Data Fig. 10b, c. C.A.M. helped to generate Extended Data Fig. 8b, d, e. J.C.W., R.A.W. and D.R.S. designed experiments, analysed the data and wrote the paper with input from the other authors. Data availability. All relevant data are available from the authors and/or are included with this Letter. Source images are available in Supplementary Fig. 1. The authors declare no competing interests. Reviewer information. Nature thanks Daniel Durocher, Michael Seidman and the other anonymous reviewer(s) for their contribution to the peer review of this work

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Accepted Version - nihms-1520599.pdf

Supplemental Material - 41586_2019_1002_Fig10_ESM.jpg

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August 22, 2023
March 5, 2024