Abstract

In response to DNA damage, tissue homoeostasis is ensured by protein networks promoting DNA repair, cell cycle arrest or apoptosis. DNA damage response signalling pathways coordinate these processes, partly by propagating gene-expression-modulating signals. DNA damage influences not only the abundance of messenger RNAs, but also their coding information through alternative splicing. Here we show that transcription-blocking DNA lesions promote chromatin displacement of late-stage spliceosomes and initiate a positive feedback loop centred on the signalling kinase ATM. We propose that initial spliceosome displacement and subsequent R-loop formation is triggered by pausing of RNA polymerase at DNA lesions. In turn, R-loops activate ATM, which signals to impede spliceosome organization further and augment ultraviolet-irradiation-triggered alternative splicing at the genome-wide level. Our findings define R-loop-dependent ATM activation by transcription-blocking lesions as an important event in the DNA damage response of non-replicating cells, and highlight a key role for spliceosome displacement in this process.

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Accessions

Primary accessions

Sequence Read Archive

Data deposits

RNA-seq data have been deposited in the Sequence Read Archive with accession number SRP053034.

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Acknowledgements

We acknowledge the Optical Imaging Center of ErasmusMC for technical support; M. Reijns and A. Jackson for the S9.6 antibody; L. Marufu for technical assistance; and N. G. J. Jaspers for intellectual input. This work was funded by the Netherlands Organization for Scientific Research (NWO) ZonMW TOP Grants 912.08.031 and 912.12.132, Horizon Zenith 935.11.042, ALW 854.11.002 and 823.02.013, the Association for International Cancer Research 10-594, European Research Council Advanced Investigator Grants 233424 and 340988, and an ErasmusMC fellowship.

Author information

Affiliations

  1. Department of Genetics, Cancer Genomics Netherlands, Erasmus University Medical Center, Rotterdam, 3015 CN, The Netherlands

    • Maria Tresini
    • , Loes Snijder
    • , Jan H. J. Hoeijmakers
    • , Wim Vermeulen
    •  & Jurgen A. Marteijn
  2. Division of Cell Biology, Netherlands Cancer Institute, Amsterdam, 1066 CX, The Netherlands

    • Daniël O. Warmerdam
    •  & René H. Medema
  3. Department of Cell Biology, Erasmus University Medical Center, Rotterdam, 3015 CN, The Netherlands

    • Petros Kolovos
    •  & Frank G. Grosveld
  4. Department of Human Genetics, Leiden University Medical Center, Leiden, 2333 ZC, The Netherlands

    • Mischa G. Vrouwe
    •  & Leon H. F. Mullenders
  5. Erasmus MC Proteomics Center, Erasmus University Medical Center, Rotterdam, 3015 CN, The Netherlands

    • Jeroen A. A. Demmers
  6. Erasmus Center for Biomics, Erasmus University Medical Center, Rotterdam, 3015 CN, The Netherlands

    • Wilfred F. J. van IJcken

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Contributions

M.T. designed the study, performed the majority of experiments, analysed the data and authored the manuscript with contributions from W.V. and J.A.M. J.A.A.D. performed the liquid-chromatography tandem mass spectrometry analysis, L.S. assisted in fractionation/immunoblotting experiments, J.A.M. performed S9.6 antibody immunofluorescence and assisted in UVC micro-irradiation experiments, D.W. and R.H.M performed RT–PCR splicing assays, P.K., F.G.G. and W.v.IJ. performed RNA-seq experiments, L.H.M and M.G.V. generated RNaseH1 constructs and cell lines. L.H.M. and J.H.J.H. provided advice. All authors reviewed and commended on the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to Maria Tresini or Wim Vermeulen or Jurgen A. Marteijn.

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DOI

https://doi.org/10.1038/nature14512

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