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Topoisomerase I suppresses genomic instability by preventing interference between replication and transcription

Nature Cell Biology volume 11, pages 1315–1324 (2009)Cite this article

A Corrigendum to this article was published on 01 November 2010

Abstract

Topoisomerase I (Top1) is a key enzyme in functioning at the interface between DNA replication, transcription and mRNA maturation. Here, we show that Top1 suppresses genomic instability in mammalian cells by preventing a conflict between transcription and DNA replication. Using DNA combing and ChIP (chromatin immunoprecipitation)-on-chip, we found that Top1-deficient cells accumulate stalled replication forks and chromosome breaks in S phase, and that breaks occur preferentially at gene-rich regions of the genome. Notably, these phenotypes were suppressed by preventing the formation of RNA–DNA hybrids (R-loops) during transcription. Moreover, these defects could be mimicked by depletion of the splicing factor ASF/SF2 (alternative splicing factor/splicing factor 2), which interacts functionally with Top1. Taken together, these data indicate that Top1 prevents replication fork collapse by suppressing the formation of R-loops in an ASF/SF2-dependent manner. We propose that interference between replication and transcription represents a major source of spontaneous replication stress, which could drive genomic instability during the early stages of tumorigenesis.

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Figure 1: Top1-deficient murine cells form DNA breaks in S phase and accumulate chromosomal aberrations.
Figure 2: Replication fork progression is impaired in the absence of Top1.
Figure 3: Analysis of sister replication-fork progression in Top1-depleted cells.
Figure 4: Inhibition of ASF/SF2 function induces fork asymmetry and chromosome breaks.
Figure 5: RNaseH1 suppresses fork asymmetry and DNA damage in Top1-deficient cells.
Figure 6: H2AX is enriched at active genes in Top1-deficient HCT116 cells.
Figure 7: Model for the role of Top1 in the coordination of DNA replication and gene expression.

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Acknowledgements

We thank M. Debatisse and M. Méchali for discussions and critical comments on the manuscript, D. Durocher for communicating data before publication, J. Piette for his initial contribution to this project and E. Schwob and the DNA combing facility of Montpellier for providing silanized coverslips. Work in the P.P. laboratory is supported by FRM (Equipe FRM), ANR, INCa and the EMBO Young Investigator Programme. Work in the J.T. laboratory is supported by ANR. A.C. is supported by the Ligue contre le cancer (Comité de l'Hérault), ARC (Association pour la Recherche contre le Cancer) and INCa. S.T., L.C. and H.T. were recipients of fellowships from ARC, EMBO and CNRS, respectively.

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  1. Arnaud Coquelle and Philippe Pasero: These authors contributed equally to this work.

Authors and Affiliations

  1. Institute of Human Genetics CNRS UPR1142, Montpellier, F-34396, France

    Sandie Tuduri, Laure Crabbé, Hélène Tourrière, Aubin Thomas & Philippe Pasero

  2. IRCM (Institut de Recherche en Cancérologie de Montpellier), INSERM U896, Université Montpellier 1, CRLC Val d'Aurelle Paul Lamarque, Montpellier, F-34298, France

    Sandie Tuduri, Charles Theillet & Arnaud Coquelle

  3. Laboratory of Molecular Pharmacology, NIH, Bethesda, 20892, MD, USA

    Chiara Conti & Yves Pommier

  4. Institute of Human Genetics, University Hospital Heidelberg, D-69120, Germany

    Heidi Holtgreve-Grez & Anna Jauch

  5. Inserm U847; CHU Montpellier, Institut de Recherche en Biothérapie, Hôpital Saint Eloi; Université Montpellier 1,

    Véronique Pantesco & John De Vos

  6. CHU Montpellier, Institut de Recherche en Biothérapie, Hôpital Saint Eloi, Montpellier, F-34000, France

    Véronique Pantesco & John De Vos

  7. Institute of Molecular Genetics, CNRS UMR5535, Montpellier, F-34293, France

    Jamal Tazi

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Contributions

A.C. and P.P. designed the project and wrote the paper; S.T., L.C., C.C. and H.T. performed most of the experiments; H.H.G. and A.J. performed the M-FISH studies; V.P. and J.D.V. performed microarray experiments. A.T. performed bioinformatic analyses; C.T. performed part of the DNA combing experiments; Y.P. and J.T. provided important experimental materials and advice and all authors contributed to interpreting the results and editing the manuscript.

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Correspondence to Arnaud Coquelle or Philippe Pasero.

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Tuduri, S., Crabbé, L., Conti, C. et al. Topoisomerase I suppresses genomic instability by preventing interference between replication and transcription. Nat Cell Biol 11, 1315–1324 (2009). https://doi.org/10.1038/ncb1984

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