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Oncogene-induced senescence is a DNA damage response triggered by DNA hyper-replication

Abstract

Early tumorigenesis is associated with the engagement of the DNA-damage checkpoint response (DDR)1,2. Cell proliferation and transformation induced by oncogene activation are restrained by cellular senescence3,4,5,6. It is unclear whether DDR activation and oncogene-induced senescence (OIS) are causally linked. Here we show that senescence, triggered by the expression of an activated oncogene (H-RasV12) in normal human cells, is a consequence of the activation of a robust DDR. Experimental inactivation of DDR abrogates OIS and promotes cell transformation. DDR and OIS are established after a hyper-replicative phase occurring immediately after oncogene expression. Senescent cells arrest with partly replicated DNA and with DNA replication origins having fired multiple times. In vivo DNA labelling and molecular DNA combing reveal that oncogene activation leads to augmented numbers of active replicons and to alterations in DNA replication fork progression. We also show that oncogene expression does not trigger a DDR in the absence of DNA replication. Last, we show that oncogene activation is associated with DDR activation in a mouse model in vivo. We propose that OIS results from the enforcement of a DDR triggered by oncogene-induced DNA hyper-replication.

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Figure 1: DNA damage accumulation and DDR activation in OIS cells.
Figure 2: DDR inactivation abolishes OIS.
Figure 3: DDR inactivation promotes cell transformation.
Figure 4: Oncogene expression leads to alterations of DNA replication.

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Acknowledgements

We thank P. Transidico for confocal analysis; IFOM Cell Biology, DNA Sequencing, Imaging and Microarrays Units for support; J. Campisi, T. D. Halazonetis, K. Helin, S. P. Jackson, Y. Shiloh, T. Mak, C. Marine, S. Pece and A. Tocchetti for sharing reagents; and M. Alcalay, B. Amati, A. Ballabeni, M. Barbacid, M. Foiani and H. Muller for discussions. F.d.A.d.F., R.M. and P.G.P. are supported by AIRC (Associazione Italiana per la Ricerca sul Cancro). R.D.M. and M.F. are SEMM (European School of Molecular Medicine) students. Author Contributions M.F. generated Fig. 4f and Supplementary Figs 4c, 7 and 17; A.C. generated Supplementary Figs S1a, b and 4a, b and helped with initial retroviral transduction experiments; S.P. and R.M. generated and analysed data in Fig. 3 and Supplementary Fig. 11; P.G. generated Fig. 4b and Supplementary Fig. 14; C.L. and P.G.N. generated and analysed data in Fig. 4g; C.S. and A.B. generated and analysed data in Fig. 4c and Supplementary Fig. 15; M.G. helped with confocal analysis; P.G.P. discussed the results; R.D.M. generated all remaining figures and contributed to experimental design; F.d.A.d.F. generated Fig. 4e, planned the project and wrote the manuscript.

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Correspondence to Fabrizio d’Adda di Fagagna.

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This file contains Supplementary Figure Legends, Supplementary Methods and additional references. (RTF 36 kb)

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This file contains Supplementary Figures 1–18. (PDF 1419 kb)

Supplementary Data

This file contains information on the genes and proteins used in this study. (DOC 22 kb)

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Di Micco, R., Fumagalli, M., Cicalese, A. et al. Oncogene-induced senescence is a DNA damage response triggered by DNA hyper-replication. Nature 444, 638–642 (2006). https://doi.org/10.1038/nature05327

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