Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

DNA end resection, homologous recombination and DNA damage checkpoint activation require CDK1

Nature volume 431, pages 1011–1017 (2004)Cite this article

Abstract

A single double-strand break (DSB) induced by HO endonuclease triggers both repair by homologous recombination and activation of the Mec1-dependent DNA damage checkpoint in budding yeast1,2,3,4,5,6. Here we report that DNA damage checkpoint activation by a DSB requires the cyclin-dependent kinase CDK1 (Cdc28) in budding yeast. CDK1 is also required for DSB-induced homologous recombination at any cell cycle stage. Inhibition of homologous recombination by using an analogue-sensitive CDK1 protein7,8 results in a compensatory increase in non-homologous end joining. CDK1 is required for efficient 5′ to 3′ resection of DSB ends and for the recruitment of both the single-stranded DNA-binding complex, RPA, and the Rad51 recombination protein. In contrast, Mre11 protein, part of the MRX complex, accumulates at unresected DSB ends. CDK1 is not required when the DNA damage checkpoint is initiated by lesions that are processed by nucleotide excision repair. Maintenance of the DSB-induced checkpoint requires continuing CDK1 activity that ensures continuing end resection. CDK1 is also important for a later step in homologous recombination, after strand invasion and before the initiation of new DNA synthesis.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: CDK1 activity is required for DSB-induced phosphorylation of checkpoint proteins in G2 cells.
Figure 2: CDK1 is required for homologous recombination.
Figure 3: CDK1 is needed for DSB resection.
Figure 4: CDK1 is required to maintain Rad53 activation in response to a DSB but not after damage by 4NQO.
Figure 5: Role of CDK1 in a late stage of MAT switching.

Similar content being viewed by others

References

  1. Lee, S. E. et al. Saccharomyces Ku70, Mre11/Rad50 and RPA proteins regulate adaptation to G2/M arrest DNA damage. Cell 94, 399–409 (1998)

    Article  CAS  Google Scholar 

  2. Toczyski, D. P., Galgoczy, D. J. & Hartwell, L. H. CDC5 and CKII control adaptation to the yeast DNA damage checkpoint. Cell 90, 1097–1106 (1997)

    Article  CAS  Google Scholar 

  3. Pellicioli, A., Lee, S. E., Lucca, C., Foiani, M. & Haber, J. E. Regulation of Saccharomyces Rad53 checkpoint kinase during adaptation from G2/M arrest. Mol. Cell 7, 293–300 (2001)

    Article  CAS  Google Scholar 

  4. Sugawara, N., Wang, X. & Haber, J. E. In vivo roles of Rad52, Rad54, and Rad55 proteins in Rad51-mediated recombination. Mol. Cell 12, 209–219 (2003)

    Article  CAS  Google Scholar 

  5. Vaze, M. B. et al. Recovery from checkpoint-mediated arrest after repair of a double-strand break requires Srs2 helicase. Mol. Cell 10, 373–385 (2002)

    Article  CAS  Google Scholar 

  6. Wang, X. & Haber, J. E. Role of Saccharomyces single-stranded DNA-binding protein RPA in the strand invasion step of double-strand break repair. PLoS Biol. 2, 104–111 (2004)

    Article  CAS  Google Scholar 

  7. Bishop, A. C. et al. A chemical switch for inhibitor-sensitive alleles of any protein kinase. Nature 407, 395–401 (2000)

    Article  ADS  CAS  Google Scholar 

  8. Ubersax, J. A. et al. Targets of the cyclin-dependent kinase Cdk1. Nature 425, 859–864 (2003)

    Article  ADS  CAS  Google Scholar 

  9. Zou, L. & Elledge, S. J. Sensing DNA damage through ATRIP recognition of RPA-ssDNA complexes. Science 300, 1542–1548 (2003)

    Article  ADS  CAS  Google Scholar 

  10. Nasmyth, K. At the heart of the budding yeast cell cycle. Trends Genet. 12, 405–412 (1996)

    Article  CAS  Google Scholar 

  11. Pellicioli, A. et al. Activation of Rad53 kinase in response to DNA damage and its effect in modulating phosphorylation of the lagging strand DNA polymerase. EMBO J. 18, 6561–6572 (1999)

    Article  CAS  Google Scholar 

  12. Naiki, T., Wakayama, T., Nakada, D., Matsumoto, K. & Sugimoto, K. Association of Rad9 with double-strand breaks through a Mec1-dependent mechanism. Mol. Cell. Biol. 24, 3277–3285 (2004)

    Article  CAS  Google Scholar 

  13. Diede, S. J. & Gottschling, D. E. Exonuclease activity is required for sequence addition and Cdc13p loading at a de novo telomere. Curr. Biol. 11, 1336–1340 (2001)

    Article  CAS  Google Scholar 

  14. Caspari, T., Murray, J. M. & Carr, A. M. Cdc2-cyclin B kinase activity links Crb2 and Rqh1-topoisomerase III. Genes Dev. 16, 1195–1208 (2002)

    Article  CAS  Google Scholar 

  15. Grenon, M., Gilbert, C. & Lowndes, N. F. Checkpoint activation in response to double-strand breaks requires the Mre11/Rad50/Xrs2 complex. Nature Cell Biol. 3, 844–847 (2001)

    Article  CAS  Google Scholar 

  16. D'Amours, D. & Jackson, S. P. The yeast Xrs2 complex functions in S phase checkpoint regulation. Genes Dev. 15, 2238–2249 (2001)

    Article  CAS  Google Scholar 

  17. Moore, J. K. & Haber, J. E. Cell cycle and genetic requirements of two pathways of nonhomologous end-joining repair of double-strand breaks in Saccharomyces cerevisiae. Mol. Cell. Biol. 16, 2164–2173 (1996)

    Article  CAS  Google Scholar 

  18. Frank-Vaillant, M. & Marcand, S. Transient stability of DNA ends allows nonhomologous end joining to precede homologous recombination. Mol. Cell 10, 1189–1199 (2002)

    Article  CAS  Google Scholar 

  19. Karathanasis, E. & Wilson, T. E. Enhancement of Saccharomyces cerevisiae end-joining efficiency by cell growth stage but not by impairment of recombination. Genetics 161, 1015–1027 (2002)

    CAS  PubMed  PubMed Central  Google Scholar 

  20. Liberi, G. et al. Srs2 DNA helicase is involved in checkpoint response and its regulation requires a functional Mec1-dependent pathway and Cdk1 activity. EMBO J. 19, 5027–5038 (2000)

    Article  CAS  Google Scholar 

  21. Sogo, J. M., Lopes, M. & Foiani, M. Fork reversal and ssDNA accumulation at stalled replication forks owing to checkpoint defects. Science 297, 599–602 (2002)

    Article  ADS  CAS  Google Scholar 

  22. Foiani, M., Liberi, G., Lucchini, G. & Plevani, P. Cell cycle-dependent phosphorylation and dephosphorylation of the yeast DNA polymerase α-primase B subunit. Mol. Cell. Biol. 15, 883–891 (1995)

    Article  CAS  Google Scholar 

  23. Lisby, M., Mortensen, U. H. & Rothstein, R. Colocalization of multiple DNA double-strand breaks at a single Rad52 repair centre. Nature Cell Biol. 5, 572–577 (2003)

    Article  CAS  Google Scholar 

  24. Rothkamm, K., Kruger, I., Thompson, L. H. & Lobrich, M. Pathways of DNA double-strand break repair during the mammalian cell cycle. Mol. Cell. Biol. 23, 5706–5715 (2003)

    Article  CAS  Google Scholar 

  25. Lee, J. & Desiderio, S. Cyclin A/CDK2 regulates V(D)J recombination by coordinating RAG-2 accumulation and DNA repair. Immunity 11, 771–781 (1999)

    Article  CAS  Google Scholar 

  26. Toh, G. W. & Lowndes, N. F. Role of the Saccharomyces cerevisiae Rad9 protein in sensing and responding to DNA damage. Biochem. Soc. Trans. 31, 242–246 (2003)

    Article  CAS  Google Scholar 

  27. Masumoto, H., Muramatsu, S., Kamimura, Y. & Araki, H. S-Cdk-dependent phosphorylation of Sld2 essential for chromosomal DNA replication in budding yeast. Nature 415, 651–655 (2002)

    Article  ADS  CAS  Google Scholar 

  28. D'Amours, D. & Jackson, S. P. The Mre11 complex: at the crossroads of DNA repair and checkpoint signalling. Nature Rev. Mol. Cell Biol. 3, 317–327 (2002)

    Article  CAS  Google Scholar 

  29. Chen, L., Trujillo, K., Ramos, W., Sung, P. & Tomkinson, A. E. Promotion of Dnl4-catalyzed DNA end-joining by the Rad50/Mre11/Xrs2 and Hdf1/Hdf2 complexes. Mol. Cell 8, 1105–1115 (2001)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The order of the co-first authors (G.I. and A.P.) and the order of the second two authors (A.B. and X.W.) were decided alphabetically. A.B. and X.W. made substantial and equal contributions to this work. We thank C. Zhang and K. Shokat (supported by NIH grant AI44009) for the cdc28-as1 allele and the 1-NMPP1 inhibitor. We note the ownership of the ASKA technology by Cellular Genomics Inc. and thank CGI for granting us the rights to perform the experiments discussed in the publication. We thank J. Petrini, J. Diffley, K. Labib, M. P. Longhese, S. Brill and the IFOM antibody facility for reagents. We are grateful to C. Lucca, E. Pandiani, M. B. Vaze, S. Francia, S. Lovett, M. Lichten and the members of our laboratories for discussions and technical support. N.M.H. was supported by the NIH. M.F. and A.P. were supported by grants from Associazione Italiana per la Ricerca sul Cancro and M.F. from Telethon-Italy and the European Community. G.I. was supported by Charles A. King Trust Postdoctoral Fellowship. G.I. is on leave from N. Copernicus University, Torun, Poland. J.E.H. was supported by the NIH and the DOE.

Author information

Author notes

  1. Xuan Wang

    Present address: Rockefeller University, 1230 York Avenue, New York, New York, 10021-6399, USA

  2. Grzegorz Ira and Achille Pellicioli: These authors contributed equally to this work

Authors and Affiliations

  1. Rosenstiel Center and Department of Biology, Brandeis University, Massachusetts, 02454-9110, Waltham, USA

    Grzegorz Ira, Xuan Wang, Debra Bressan & James E. Haber

  2. F.I.R.C. Institute of Molecular Oncology Foundation, Via Adamello 16, 20139, Milano, Italy

    Achille Pellicioli, Alitukiriza Balijja, Simona Fiorani, Walter Carotenuto, Giordano Liberi & Marco Foiani

  3. Dipartimento di Scienze Biomolecolari e Biotecnologie, Universitá degli Studi di Milano, Italy

    Achille Pellicioli, Alitukiriza Balijja, Simona Fiorani, Walter Carotenuto, Giordano Liberi & Marco Foiani

  4. Department of Biochemistry and Cell Biology, SUNY Stony Brook, Stony Brook, New York, 11794-5215, USA

    Lihong Wan & Nancy M. Hollingsworth

Authors
  1. Grzegorz Ira
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Achille Pellicioli
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Alitukiriza Balijja
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xuan Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Simona Fiorani
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Walter Carotenuto
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Giordano Liberi
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Debra Bressan
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Lihong Wan
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Nancy M. Hollingsworth
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. James E. Haber
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Marco Foiani
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to James E. Haber or Marco Foiani.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Supplementary Figure

Graph and gel scans. (PDF 174 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ira, G., Pellicioli, A., Balijja, A. et al. DNA end resection, homologous recombination and DNA damage checkpoint activation require CDK1. Nature 431, 1011–1017 (2004). https://doi.org/10.1038/nature02964

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature02964

This article is cited by

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing