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NONO regulates the intra-S-phase checkpoint in response to UV radiation

Oncogene volume 35, pages 567–576 (2016)Cite this article

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Abstract

The main risk factor for skin cancer is ultraviolet (UV) exposure, which causes DNA damage. Cells respond to UV-induced DNA damage by activating the intra-S-phase checkpoint, which prevents replication fork collapse, late origin firing and stabilizes fragile sites. Recently, the 54-kDa multifunctional protein NONO was found to be involved in the non-homologous end-joining DNA repair process and in poly ADP-ribose polymerase 1 activation. Interestingly, NONO is mutated in several tumour types and emerged as a crucial factor underlying both melanoma development and progression. Therefore, we set out to evaluate whether NONO could be involved in the DNA-damage response to UV radiations. We generated NONO-silenced HeLa cell clones and found that lack of NONO decreased cell growth rate. Then, we challenged NONO-silenced cells with exposure to UV radiations and found that NONO-silenced cells, compared with control cells, continued to synthesize DNA, failed to block new origin firing and impaired CHK1S345 phosphorylation showing a defective checkpoint activation. Consistently, NONO is present at the sites of UV-induced DNA damage where it localizes to RAD9 foci. To position NONO in the DNA-damage response cascade, we analysed the loading onto chromatin of various intra-S-phase checkpoint mediators and found that NONO favours the loading of topoisomerase II-binding protein 1 acting upstream of the ATM and Rad3-related kinase activity. Strikingly, re-expression of NONO, through an sh-resistant mRNA, rescued CHK1S345 phosphorylation in NONO-silenced cells. Interestingly, NONO silencing affected cell response to UV radiations also in a melanoma cell line. Overall, our data uncover a new role for NONO in mediating the cellular response to UV-induced DNA damage.

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Acknowledgements

We are thankful to the Sbarro Health Research Organization (http://www.shro.org), the Human Health Foundation (http://www.hhfonlus.org), the Commonwealth of Pennsylvania and the Associazione Italiana per la Ricerca sul Cancro (AIRC IG 15690) for their support. We are grateful to Pasquale Barba and Laura Pisapia (Consiglio Nazionale delle Ricerche-Institute of Genetics and Biophysics, Naples, Italy) for the FACS analyses. We also thank Larry M Karnitz, Division of Oncology Research, Mayo Clinic, Rochester, MN, USA for his help in sharing reagents and Marco G Paggi, Regina Elena National Cancer Institute, Rome, Italy for the M14 cell line. Prof Giordano is the director of research line 3 at CROM, Istituto Nazionale Tumori 'Fondazione G. Pascale'- IRCCS, Naples, Italy.

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Authors and Affiliations

  1. CROM, Istituto Nazionale Tumori “Fondazione G. Pascale”- IRCCS,, Naples, Italy

    L Alfano, C Costa, A Altieri & F Pentimalli

  2. Department of Medicine, Surgery and Neuroscience, University of Siena and Istituto Toscano Tumori (ITT), Siena, Italy

    A Caporaso, P Indovina & A Giordano

  3. Sbarro Institute for Cancer Research and Molecular Medicine and Center for Biotechnology, College of Science and Technology, Temple University, Philadelphia, PA, USA

    P Indovina, M Macaluso & A Giordano

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  1. L Alfano
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Correspondence to A Giordano or F Pentimalli.

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Alfano, L., Costa, C., Caporaso, A. et al. NONO regulates the intra-S-phase checkpoint in response to UV radiation. Oncogene 35, 567–576 (2016). https://doi.org/10.1038/onc.2015.107

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