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DNA damage induces Chk1-dependent threonine-160 phosphorylation and activation of Cdk2

Oncogene volume 29, pages 616–624 (2010)Cite this article

Abstract

Abnormal centrosome numbers arise in tumours and can cause multipolar mitoses and genome instability. Cdk2 controls normal centrosome duplication, but Chk1-dependent centrosome amplification also occurs after DNA damage. We investigated the involvement of cyclin-dependent kinases (Cdks) in DNA damage-induced centrosome amplification using cells lacking either Cdk2, or both Cdk1 and Cdk2 activity. Cdk2−/− DT40 cells showed robust centrosome amplification after ionizing radiation (IR), whereas Cdk1-deficient Cdk2−/− cells showed no centrosome amplification, demonstrating that Cdk1 can substitute for Cdk2 in this pathway. Surprisingly, we found that Cdk2 activity was upregulated by IR in wild-type but not in Chk1−/− DT40 cells. Cdk2 upregulation also occurred in HeLa cells after IR treatment. Chk1-dependent Cdk2 induction was not accompanied by increased levels of Cdk1, Cdk2, cyclin A or cyclin E, but activating T160 phosphorylation of Cdk2 increased after IR. Moreover, Cdk2 overexpression restored IR-induced centrosome amplification in Cdk1-deficient Cdk2−/− cells, but T160A mutation blocked this rescue. Our data suggest that Chk1 signalling causes centrosome amplification after IR by upregulating Cdk2 activity through activating phosphorylation.

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References

  • Aleem E, Kiyokawa H, Kaldis P . (2005). Cdc2-cyclin E complexes regulate the G1/S phase transition. Nat Cell Biol 7: 831–836.

    Article  CAS  PubMed  Google Scholar 

  • Balczon RC . (2001). Overexpression of cyclin A in human HeLa cells induces detachment of kinetochores and spindle pole/centrosome overproduction. Chromosoma 110: 381–392.

    Article  CAS  PubMed  Google Scholar 

  • Beamish H, Williams R, Chen P, Lavin MF . (1996). Defect in multiple cell cycle checkpoints in ataxia-telangiectasia postirradiation. J Biol Chem 271: 20486–20493.

    Article  CAS  PubMed  Google Scholar 

  • Berthet C, Aleem E, Coppola V, Tessarollo L, Kaldis P . (2003). Cdk2 knockout mice are viable. Curr Biol 13: 1775–1785.

    Article  CAS  PubMed  Google Scholar 

  • Berthet C, Kaldis P . (2007). Cell-specific responses to loss of cyclin-dependent kinases. Oncogene 26: 4469–4477.

    Article  CAS  PubMed  Google Scholar 

  • Bettencourt-Dias M, Glover DM . (2007). Centrosome biogenesis and function: centrosomics brings new understanding. Nat Rev Mol Cell Biol 8: 451–463.

    Article  CAS  PubMed  Google Scholar 

  • Bourke E, Dodson H, Merdes A, Cuffe L, Zachos G, Walker M et al. (2007). DNA damage induces Chk1-dependent centrosome amplification. EMBO Rep 8: 603–609.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cannell EJ, Farrell PJ, Sinclair AJ . (1998). Cell cycle arrest following exposure of EBV-immortalised B-cells to gamma irradiation correlates with inhibition of cdk2 activity. FEBS Lett 439: 297–301.

    Article  CAS  PubMed  Google Scholar 

  • Chen MS, Ryan CE, Piwnica-Worms H . (2003). Chk1 kinase negatively regulates mitotic function of Cdc25A phosphatase through 14-3-3 binding. Mol Cell Biol 23: 7488–7497.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dodson H, Bourke E, Jeffers LJ, Vagnarelli P, Sonoda E, Takeda S et al. (2004). Centrosome amplification induced by DNA damage occurs during a prolonged G2 phase and involves ATM. EMBO J 23: 3864–3873.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Dodson H, Wheatley SP, Morrison CG . (2007). Involvement of centrosome amplification in radiation-induced mitotic catastrophe. Cell Cycle 6: 364–370.

    Article  CAS  PubMed  Google Scholar 

  • Doxsey S, McCollum D, Theurkauf W . (2005). Centrosomes in cellular regulation. Annu Rev Cell Dev Biol 21: 411–434.

    Article  CAS  PubMed  Google Scholar 

  • Duensing A, Liu Y, Tseng M, Malumbres M, Barbacid M, Duensing S . (2006). Cyclin-dependent kinase 2 is dispensable for normal centrosome duplication but required for oncogene-induced centrosome overduplication. Oncogene 25: 2943–2949.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fukasawa K . (2007). Oncogenes and tumour suppressors take on centrosomes. Nat Rev Cancer 7: 911–924.

    Article  CAS  PubMed  Google Scholar 

  • Fukasawa K, Choi T, Kuriyama R, Rulong S, Vande Woude GF . (1996). Abnormal centrosome amplification in the absence of p53. Science 271: 1744–1747.

    Article  CAS  PubMed  Google Scholar 

  • Ganem NJ, Godinho SA, Pellman D . (2009). A mechanism linking extra centrosomes to chromosomal instability. Nature 460: 278–282.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Hanashiro K, Kanai M, Geng Y, Sicinski P, Fukasawa K . (2008). Roles of cyclins A and E in induction of centrosome amplification in p53-compromised cells. Oncogene 27: 5288–5302.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • He G, Siddik ZH, Huang Z, Wang R, Koomen J, Kobayashi R et al. (2005). Induction of p21 by p53 following DNA damage inhibits both Cdk4 and Cdk2 activities. Oncogene 24: 2929–2943.

    Article  CAS  PubMed  Google Scholar 

  • Hinchcliffe EH, Li C, Thompson EA, Maller JL, Sluder G . (1999). Requirement of Cdk2-cyclin E activity for repeated centrosome reproduction in Xenopus egg extracts. Science 283: 851–854.

    Article  CAS  PubMed  Google Scholar 

  • Hochegger H, Dejsuphong D, Sonoda E, Saberi A, Rajendra E, Kirk J et al. (2007). An essential role for Cdk1 in S phase control is revealed via chemical genetics in vertebrate cells. J Cell Biol 178: 257–268.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kawamura K, Izumi H, Ma Z, Ikeda R, Moriyama M, Tanaka T et al. (2004). Induction of centrosome amplification and chromosome instability in human bladder cancer cells by p53 mutation and cyclin E overexpression. Cancer Res 64: 4800–4809.

    Article  CAS  PubMed  Google Scholar 

  • Kramer A, Mailand N, Lukas C, Syljuasen RG, Wilkinson CJ, Nigg EA et al. (2004). Centrosome-associated Chk1 prevents premature activation of cyclin-B-Cdk1 kinase. Nat Cell Biol 6: 884–891.

    Article  PubMed  Google Scholar 

  • Lacey KR, Jackson PK, Stearns T . (1999). Cyclin-dependent kinase control of centrosome duplication. Proc Natl Acad Sci USA 96: 2817–2822.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Mailand N, Falck J, Lukas C, Syljuasen RG, Welcker M, Bartek J et al. (2000). Rapid destruction of human Cdc25A in response to DNA damage. Science 288: 1425–1429.

    Article  CAS  PubMed  Google Scholar 

  • Mailand N, Podtelejnikov AV, Groth A, Mann M, Bartek J, Lukas J . (2002). Regulation of G(2)/M events by Cdc25A through phosphorylation-dependent modulation of its stability. EMBO J 21: 5911–5920.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Matsumoto Y, Hayashi K, Nishida E . (1999). Cyclin-dependent kinase 2 (Cdk2) is required for centrosome duplication in mammalian cells. Curr Biol 9: 429–432.

    Article  CAS  PubMed  Google Scholar 

  • Meraldi P, Lukas J, Fry AM, Bartek J, Nigg EA . (1999). Centrosome duplication in mammalian somatic cells requires E2F and Cdk2-cyclin A. Nat Cell Biol 1: 88–93.

    Article  CAS  PubMed  Google Scholar 

  • Muller-Tidow C, Ji P, Diederichs S, Potratz J, Baumer N, Kohler G et al. (2004). The cyclin A1-CDK2 complex regulates DNA double-strand break repair. Mol Cell Biol 24: 8917–8928.

    Article  PubMed  PubMed Central  Google Scholar 

  • Mussman JG, Horn HF, Carroll PE, Okuda M, Tarapore P, Donehower LA et al. (2000). Synergistic induction of centrosome hyperamplification by loss of p53 and cyclin E overexpression. Oncogene 19: 1635–1646.

    Article  CAS  PubMed  Google Scholar 

  • Nigg EA . (2006). Origins and consequences of centrosome aberrations in human cancers. Int J Cancer 119: 2717–2723.

    Article  CAS  PubMed  Google Scholar 

  • Ortega S, Prieto I, Odajima J, Martin A, Dubus P, Sotillo R et al. (2003). Cyclin-dependent kinase 2 is essential for meiosis but not for mitotic cell division in mice. Nat Genet 35: 25–31.

    Article  CAS  PubMed  Google Scholar 

  • Prosser SL, Straatman KR, Fry AM . (2009). Molecular dissection of the centrosome overduplication pathway in S-phase arrested cells. Mol Cell Biol 29: 1760–1773.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Saladino C, Bourke E, Conroy P, Morrison CG . (2009). Centriole separation in DNA damage-induced centrosome amplification. Environ Mol Mutagen 50: 725–732.

    Article  CAS  PubMed  Google Scholar 

  • Sampath D, Shi Z, Plunkett W . (2002). Inhibition of cyclin-dependent kinase 2 by the Chk1-Cdc25A pathway during the S-phase checkpoint activated by fludarabine: dysregulation by 7-hydroxystaurosporine. Mol Pharmacol 62: 680–688.

    Article  CAS  PubMed  Google Scholar 

  • Sanchez Y, Wong C, Thoma RS, Richman R, Wu Z, Piwnica-Worms H et al. (1997). Conservation of the Chk1 checkpoint pathway in mammals: linkage of DNA damage to Cdk regulation through Cdc25. Science 277: 1497–1501.

    Article  CAS  PubMed  Google Scholar 

  • Sato N, Mizumoto K, Nakamura M, Tanaka M . (2000). Radiation-induced centrosome overduplication and multiple mitotic spindles in human tumor cells. Exp Cell Res 255: 321–326.

    Article  CAS  PubMed  Google Scholar 

  • Satyanarayana A, Hilton MB, Kaldis P . (2008). p21 Inhibits Cdk1 in the absence of Cdk2 to maintain the G1/S phase DNA damage checkpoint. Mol Biol Cell 19: 65–77.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shimuta K, Nakajo N, Uto K, Hayano Y, Okazaki K, Sagata N . (2002). Chk1 is activated transiently and targets Cdc25A for degradation at the Xenopus midblastula transition. EMBO J 21: 3694–3703.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shreeram S, Hee WK, Bulavin DV . (2008). Cdc25A serine 123 phosphorylation couples centrosome duplication with DNA replication and regulates tumorigenesis. Mol Cell Biol 28: 7442–7450.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Spruck CH, Won KA, Reed SI . (1999). Deregulated cyclin E induces chromosome instability. Nature 401: 297–300.

    Article  CAS  PubMed  Google Scholar 

  • Tarapore P, Fukasawa K . (2002). Loss of p53 and centrosome hyperamplification. Oncogene 21: 6234–6240.

    Article  CAS  PubMed  Google Scholar 

  • Tsou MF, Stearns T . (2006). Mechanism limiting centrosome duplication to once per cell cycle. Nature 442: 947–951.

    Article  CAS  PubMed  Google Scholar 

  • Uto K, Inoue D, Shimuta K, Nakajo N, Sagata N . (2004). Chk1, but not Chk2, inhibits Cdc25 phosphatases by a novel common mechanism. EMBO J 23: 3386–3396.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wong C, Stearns T . (2003). Centrosome number is controlled by a centrosome-intrinsic block to reduplication. Nat Cell Biol 5: 539–544.

    Article  CAS  PubMed  Google Scholar 

  • Yamazoe M, Sonoda E, Hochegger H, Takeda S . (2004). Reverse genetic studies of the DNA damage response in the chicken B lymphocyte line DT40. DNA Repair (Amst) 3: 1175–1185.

    Article  CAS  Google Scholar 

  • Zachos G, Rainey MD, Gillespie DA . (2003). Chk1-deficient tumour cells are viable but exhibit multiple checkpoint and survival defects. EMBO J 22: 713–723.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Zhao H, Watkins JL, Piwnica-Worms H . (2002). Disruption of the checkpoint kinase 1/cell division cycle 25A pathway abrogates ionizing radiation-induced S and G2 checkpoints. Proc Natl Acad Sci USA 99: 14795–14800.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

This work was supported by Science Foundation Ireland Principal Investigator award 08/IN.1/B1029 and by Health Research Board project grant RP/2006/36. We thank Helen Dodson for helping us with the initial Cdk kinase assays and Corrado Santocanale for critical reading of the paper.

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Authors and Affiliations

  1. Centre for Chromosome Biology, School of Natural Sciences, National University of Ireland Galway, Galway, Ireland

    E Bourke, J A L Brown & C G Morrison

  2. Department of Radiation Genetics, Kyoto University Medical School, Kyoto, Japan

    S Takeda

  3. Genome Damage and Stability Centre, Science Park Road, University of Sussex, Falmer, Brighton, UK

    H Hochegger

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  1. E Bourke
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Correspondence to C G Morrison.

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Supplementary Information accompanies the paper on the Oncogene website (http://www.nature.com/onc)

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Bourke, E., Brown, J., Takeda, S. et al. DNA damage induces Chk1-dependent threonine-160 phosphorylation and activation of Cdk2. Oncogene 29, 616–624 (2010). https://doi.org/10.1038/onc.2009.340

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