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:

MDC1 is required for the intra-S-phase DNA damage checkpoint

Nature volume 421, pages 952–956 (2003)Cite this article

Abstract

MRE11, RAD50 and NBS1 form a highly conserved protein complex (the MRE11 complex) that is involved in the detection, signalling and repair of DNA damage1. We identify MDC1 (KIAA0170/NFBD1), a protein that contains a forkhead-associated (FHA) domain and two BRCA1 carboxy-terminal (BRCT) domains, as a binding partner for the MRE11 complex. We show that, in response to ionizing radiation, MDC1 is hyperphosphorylated in an ATM-dependent manner, and rapidly relocalizes to nuclear foci that also contain the MRE11 complex, phosphorylated histone H2AX and 53BP1. Downregulation of MDC1 expression by small interfering RNA yields a radio-resistant DNA synthesis (RDS) phenotype and prevents ionizing radiation-induced focus formation by the MRE11 complex. However, downregulation of MDC1 does not abolish the ionizing radiation-induced phosphorylation of NBS1, CHK2 and SMC1, or the degradation of CDC25A. Furthermore, we show that overexpression of the MDC1 FHA domain interferes with focus formation by MDC1 itself and by the MRE11 complex, and induces an RDS phenotype. These findings reveal that MDC1-mediated focus formation by the MRE11 complex at sites of DNA damage is crucial for the efficient activation of the intra-S-phase checkpoint.

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: Association of MDC1 with the MRE11 complex.
Figure 2: MDC1 responds to ionizing radiation.
Figure 3: MDC1 focus formation does not require ATM, ATR, DNA-PK or NBS1.
Figure 5: MDC1 functions in the S-phase checkpoint.
Figure 4: MDC1 is required to recruit the MRE11 complex to foci.

Similar content being viewed by others

References

  1. 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 

  2. Gatei, M. et al. ATM-dependent phosphorylation of nibrin in response to radiation exposure. Nature Genet. 25, 115–119 (2000)

    Article  CAS  Google Scholar 

  3. Lim, D. S. et al. ATM phosphorylates p95/nbs1 in an S-phase checkpoint pathway. Nature 404, 613–617 (2000)

    Article  ADS  CAS  Google Scholar 

  4. Wu, X. et al. ATM phosphorylation of Nijmegen breakage syndrome protein is required in a DNA damage response. Nature 405, 477–482 (2000)

    Article  ADS  CAS  Google Scholar 

  5. Zhao, S. et al. Functional link between ataxia–telangiectasia and Nijmegen breakage syndrome gene products. Nature 405, 473–477 (2000)

    Article  ADS  CAS  Google Scholar 

  6. Carney, J. P. et al. The hMre11/hRad50 protein complex and Nijmegen breakage syndrome: Linkage of double-strand break repair to the cellular DNA damage response. Cell 93, 477–486 (1998)

    Article  CAS  Google Scholar 

  7. Matsuura, S. et al. Positional cloning of the gene for Nijmegen breakage syndrome. Nature Genet. 19, 179–181 (1998)

    Article  CAS  Google Scholar 

  8. Stewart, G. S. et al. The DNA double-strand break repair gene hMRE11 is mutated in individuals with an ataxia–telangiectasia-like disorder. Cell 99, 577–587 (1999)

    Article  CAS  Google Scholar 

  9. Savitsky, K. et al. A single ataxia telangiectasia gene with a product similar to PI-3 kinase. Science 268, 1749–1753 (1995)

    Article  ADS  CAS  Google Scholar 

  10. Varon, R. et al. Nibrin, a novel DNA double-strand break repair protein, is mutated in Nijmegen breakage syndrome. Cell 93, 467–476 (1998)

    Article  CAS  Google Scholar 

  11. Michelson, R. J. & Weinert, T. Closing the gaps among a web of DNA repair disorders. BioEssays 22, 966–969 (2000)

    Article  CAS  Google Scholar 

  12. Shiloh, Y. Ataxia–telangiectasia and the Nijmegen breakage syndrome: Related disorders but genes apart. Annu. Rev. Genet. 31, 635–662 (1997)

    Article  CAS  Google Scholar 

  13. Falck, J., Petrini, J. H., Williams, B. R., Lukas, J. & Bartek, J. The DNA damage-dependent intra-S phase checkpoint is regulated by parallel pathways. Nature Genet. 30, 290–294 (2002)

    Article  Google Scholar 

  14. Nagase, T., Seki, N., Ishikawa, K., Tanaka, A. & Nomura, N. Prediction of the coding sequences of unidentified human genes. V. The coding sequences of 40 new genes (KIAA0161–KIAA0200) deduced by analysis of cDNA clones from human cell line KG-1. DNA Res. 3, 17–24 (1996)

    Article  CAS  Google Scholar 

  15. Durocher, D. & Jackson, S. P. The FHA domain. FEBS Lett. 513, 58–66 (2002)

    Article  CAS  Google Scholar 

  16. Bork, P. et al. A superfamily of conserved domains in DNA damage-responsive cell cycle checkpoint proteins. FASEB J. 11, 68–76 (1997)

    Article  CAS  Google Scholar 

  17. Abraham, R. T. Cell cycle checkpoint signaling through the ATM and ATR kinases. Genes Dev. 15, 2177–2196 (2001)

    Article  CAS  Google Scholar 

  18. Maser, R. S., Monsen, K. J., Nelms, B. E. & Petrini, J. H. hMre11 and hRad50 nuclear foci are induced during the normal cellular response to DNA double-strand breaks. Mol. Cell. Biol. 17, 6087–6096 (1997)

    Article  CAS  Google Scholar 

  19. Schultz, L. B., Chehab, N. H., Malikzay, A. & Halazonetis, T. D. p53 binding protein 1 (53BP1) is an early participant in the cellular response to DNA double-strand breaks. J. Cell Biol. 151, 1381–1390 (2000)

    Article  CAS  Google Scholar 

  20. Paull, T. T. et al. A critical role for histone H2AX in recruitment of repair factors to nuclear foci after DNA damage. Curr. Biol. 10, 886–895 (2000)

    Article  CAS  Google Scholar 

  21. Rogakou, E. P., Boon, C., Redon, C. & Bonner, W. M. Megabase chromatin domains involved in DNA double-strand breaks in vivo. J. Cell Biol. 146, 905–916 (1999)

    Article  CAS  Google Scholar 

  22. Burma, S., Chen, B. P., Murphy, M., Kurimasa, A. & Chen, D. J. ATM phosphorylates histone H2AX in response to DNA double-strand breaks. J. Biol. Chem. 276, 42462–42467 (2001)

    Article  CAS  Google Scholar 

  23. Sarkaria, J. N. et al. Inhibition of phosphoinositide 3-kinase related kinases by the radiosensitizing agent wortmannin. Cancer Res. 58, 4375–4382 (1998)

    CAS  PubMed  Google Scholar 

  24. Elbashir, S. M. et al. Duplexes of 21-nucleotide RNAs mediate RNA interference in cultured mammalian cells. Nature 411, 494–498 (2001)

    Article  ADS  CAS  Google Scholar 

  25. Li, J. et al. Structural and functional versatility of the FHA domain in DNA-damage signaling by the tumour suppressor kinase Chk2. Mol. Cell 9, 1045–1054 (2002)

    Article  CAS  Google Scholar 

  26. Falck, J., Mailand, N., Syljuasen, R. G., Bartek, J. & Lukas, J. The ATM–Chk2–Cdc25A checkpoint pathway guards against radioresistant DNA synthesis. Nature 410, 842–847 (2001)

    Article  ADS  CAS  Google Scholar 

  27. Yazdi, P. T. et al. SMC1 is a downstream effector in the ATM/NBS1 branch of the human S-phase checkpoint. Genes Dev. 16, 571–582 (2002)

    Article  CAS  Google Scholar 

  28. Kim, S. T., Xu, B. & Kastan, M. B. Involvement of the cohesin protein, Smc1, in Atm-dependent and independent responses to DNA damage. Genes Dev. 16, 560–570 (2002)

    Article  CAS  Google Scholar 

  29. Maser, R. S. et al. Mre11 complex and DNA replication: Linkage to E2F and sites of DNA synthesis. Mol. Cell Biol. 21, 6006–6016 (2001)

    Article  CAS  Google Scholar 

  30. Ozaki, T. et al. NFBD1/KIAA0170 is a novel nuclear transcriptional transactivator with BRCT domain. DNA Cell Biol. 19, 475–485 (2000)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank Y. Shiloh for AT complemented cells, T. D. Halazonetis for anti-53BP1 antibodies, J. Petrini for anti-MRE11 and anti-NBS1 antibodies, and M. Kastan and K. K. Khanna for anti-NBS1-S343(P) antibodies. We thank S.P.J. laboratory members for helpful discussions and critical reading of the manuscript, especially J. Bradbury, F. d'Adda di Fagagna, M. Grenon, J. Rouse and V. Smits. We thank S. Elledge for sharing unpublished results. This study was supported by grants from the Association for International Cancer Research, the A-T Medical Research Trust and the Swiss National Foundation. Research in the S.P.J. laboratory is funded by Cancer Research UK.

Author information

Author notes

  1. Michal Goldberg, Manuel Stucki, Dinah Rahman and Darryl Pappin: These authors contributed equally to this work

Authors and Affiliations

  1. The Wellcome Trust/Cancer Research UK Institute of Cancer and Developmental Biology and Department of Zoology, University of Cambridge, CB2 1QR, Cambridge, UK

    Michal Goldberg, Manuel Stucki, Damien D'Amours & Stephen P. Jackson

  2. Institute of Cancer Biology, Danish Cancer Society, Strandboulevarden 49, DK-2100, Copenhagen, Denmark

    Jacob Falck & Jiri Bartek

  3. Department of Proteomics, Imperial College School of Medicine, Hammersmith Campus, W12 0NN, London, UK

    Dinah Rahman & Darryl Pappin

Authors
  1. Michal Goldberg
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Manuel Stucki
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Jacob Falck
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Damien D'Amours
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Dinah Rahman
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Darryl Pappin
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Jiri Bartek
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Stephen P. Jackson
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Stephen P. Jackson.

Ethics declarations

Competing interests

S. Jackson founded KuDOS Pharmaceuticals Ltd and is chief scientific officer of the company. KuDOS is using knowledge of DNA repair pathways to discover and develop new therapies for cancer and other human diseases.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Goldberg, M., Stucki, M., Falck, J. et al. MDC1 is required for the intra-S-phase DNA damage checkpoint. Nature 421, 952–956 (2003). https://doi.org/10.1038/nature01445

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature01445

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