Abstract

Error-free repair of DNA double-strand breaks (DSBs) is achieved by homologous recombination (HR), and BRCA1 is an important factor for this repair pathway1. In the absence of BRCA1-mediated HR, the administration of PARP inhibitors induces synthetic lethality of tumour cells of patients with breast or ovarian cancers2,3. Despite the benefit of this tailored therapy, drug resistance can occur by HR restoration4. Genetic reversion of BRCA1-inactivating mutations can be the underlying mechanism of drug resistance, but this does not explain resistance in all cases5. In particular, little is known about BRCA1-independent restoration of HR. Here we show that loss of REV7 (also known as MAD2L2) in mouse and human cell lines re-establishes CTIP-dependent end resection of DSBs in BRCA1-deficient cells, leading to HR restoration and PARP inhibitor resistance, which is reversed by ATM kinase inhibition. REV7 is recruited to DSBs in a manner dependent on the H2AX–MDC1–RNF8–RNF168–53BP1 chromatin pathway, and seems to block HR and promote end joining in addition to its regulatory role in DNA damage tolerance6. Finally, we establish that REV7 blocks DSB resection to promote non-homologous end-joining during immunoglobulin class switch recombination. Our results reveal an unexpected crucial function of REV7 downstream of 53BP1 in coordinating pathological DSB repair pathway choices in BRCA1-deficient cells.

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Acknowledgements

We thank B. Gerritsen, P. Halonen, B. Morris, T. Halazonetis and O. Kallioniemi for advice on the DDR shRNA library, A. Gasparini and G. Borst for their assistance with the cone beam micro-irradiator, R. Kanaar for his RAD51 antibody, J. Jacobs for the pMSCV-GFP vector, and M. O’Connor for olaparib and AZD2461. This work was supported by the Netherlands Organization for Scientific Research (NWO-Toptalent to J.E.J. and NWO-VIDI to S.R.), the Dutch Cancer Society, CTMM Breast Care, the Swiss National Science Foundation, and the European Union (EU) FP6 Integrated Project CHEMORES and FP7 Project DDResponse. Work in J.R.C.’s group is funded by the Wellcome Trust. The work in the J.B.’s laboratories was funded by the Danish Cancer Society, the Danish Council for Independent Research, the Lundbeck Foundation and the Czech National Program of Sustainability. S.J.B. is funded by Cancer Research UK and an ERC Advanced Investigator Grant (RecMitMei) and is a Royal Society Wolfson Research Merit Award Holder.

Author information

Author notes

    • J. Ross Chapman
    •  & Inger Brandsma

    These authors contributed equally to this work.

Affiliations

  1. Division of Molecular Oncology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Guotai Xu
    • , Ewa Gogola
    • , Marco Barazas
    • , Janneke E. Jaspers
    • , Ariena Kersbergen
    • , Wendy Sol
    • , Piet Borst
    •  & Sven Rottenberg
  2. The Wellcome Trust Centre for Human Genetics, Roosevelt Drive, Oxford OX3 7BN, UK

    • J. Ross Chapman
    •  & Ahmed Salman
  3. Department of Genetics, Erasmus, University Medical Center, 3000 CA Rotterdam, The Netherlands

    • Inger Brandsma
    •  & Dik C. van Gent
  4. Department of Experimental Radiation Oncology, University of Texas MD Anderson Cancer Center, Houston, Texas 77030, USA

    • Jingsong Yuan
    •  & Junjie Chen
  5. Institute of Molecular and Translational Medicine, Faculty of Medicine and Dentistry, Palacky University, 779 00 Olomouc, Czech Republic

    • Martin Mistrik
    •  & Jiri Bartek
  6. Division of Molecular Pathology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Peter Bouwman
    • , Mark Pieterse
    • , Philip C. Schouten
    •  & Jos Jonkers
  7. Danish Cancer Society Research Center, 2100 Copenhagen, Denmark

    • Jirina Bartkova
    • , Kenji Watanabe
    •  & Jiri Bartek
  8. Division of Cell Biology, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Daniël Warmerdam
    • , Bram van den Broek
    •  & Kees Jalink
  9. Protein Facility, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Patrick H. N. Celie
  10. Deep Sequencing Core Facility, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Marja Nieuwland
    •  & Iris de Rink
  11. Division of Molecular Carcinogenesis, The Netherlands Cancer Institute, Plesmanlaan 121, 1066CX Amsterdam, The Netherlands

    • Jorma de Ronde
  12. DNA Damage Response Laboratory, London Research Institute, Cancer Research UK, Clare Hall, South Mimms, Hertfordshire EN6 3LD, UK

    • Simon J. Boulton
  13. Institute of Animal Pathology, Vetsuisse Faculty, University of Bern, Laengassstrasse 122, 3012 Bern, Switzerland

    • Sven Rottenberg

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Contributions

G.X. and S.R. designed the study, performed experiments and wrote the manuscript; I.B. and D.C.v.G. designed and performed the RPA foci analysis; J.R.C., A.S. and S.J.B. performed and planned the CSR assay and RIF1-associated data; J.C. and J.Y. designed and performed the experiments using MEFs; J.E.J., A.K. and W.S. assisted with the mouse intervention studies; E.G. established the in vivo RAD51 analysis that K.J. and B.v.d.B. quantified; P.H.N.C. designed and M.B. helped in generating the REV7 mutants; Pe.B., M.P. and J.J. helped in designing the shRNA screen and performed experiments using mES cells; M.M. performed the laser stripe assays, and K.W. performed co-immunoprecipitations. D.W. helped to visualize GFP–REV7 recruitment; M.N., I.d.R. and J.d.R. carried out the RNA sequencing (RNA-seq) data analysis, Jirina B. established, carried out and evaluated the REV7 immunohistochemistry, P.C.S. helped with the analysis of the immunohistochemistry data, Jiri B. and Pi.B. advised on experiments and manuscript revisions, and all authors discussed and approved the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to Sven Rottenberg.

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https://doi.org/10.1038/nature14328

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