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:

Roles of ATM and NBS1 in chromatin structure modulation and DNA double-strand break repair

Nature Cell Biology volume 9, pages 683–690 (2007)Cite this article

Abstract

We developed a novel system to create DNA double-strand breaks (DSBs) at defined endogenous sites in the human genome, and used this system to detect protein recruitment and loss at and around these breaks by chromatin immunoprecipitation (ChIP). The detection of human ATM protein at site-specific DSBs required functional NBS1 protein, ATM kinase activity and ATM autophosphorylation on Ser 1981. DSB formation led to the localized disruption of nucleosomes, a process that depended on both functional NBS1 and ATM. These two proteins were also required for efficient recruitment of the repair cofactor XRCC4 to DSBs, and for efficient DSB repair. These results demonstrate the functional importance of ATM kinase activity and phosphorylation in the response to DSBs, and support a model in which ordered chromatin structure changes that occur after DNA breakage depend on functional NBS1 and ATM, and facilitate DNA DSB repair.

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

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

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

Figure 1: Characterization of I-PpoI-mediated DSB formation and damage response.
Figure 2: ATM dimers are not detected at DSB sites.
Figure 3: Detection, chromatin modulation and repair of a site-specific DSB.
Figure 4: Histone loss from DSB sites and DSB repair are dependent on NBS1 and ATM.
Figure 5: Distribution of damage response proteins around a site-specific DSB.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. 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 

  2. Rudin, N. & Haber, J. E. Efficient repair of HO-induced chromosomal breaks in Saccharomyces cerevisiae by recombination between flanking homologous sequences. Mol. Cell Biol. 8, 3918–3928 (1988).

    Article  CAS  Google Scholar 

  3. Tsukuda, T., Fleming, A. B., Nickoloff, J. A. & Osley, M. A. Chromatin remodelling at a DNA double-strand break site in Saccharomyces cerevisiae. Nature 438, 379–383 (2005).

    Article  CAS  Google Scholar 

  4. Lisby, M., Barlow, J. H., Burgess, R. C. & Rothstein, R. Choreography of the DNA damage response: spatiotemporal relationships among checkpoint and repair proteins. Cell 118, 699–713 (2004).

    Article  CAS  Google Scholar 

  5. Richardson, C. & Jasin, M. Frequent chromosomal translocations induced by DNA double-strand breaks. Nature 405, 697–700 (2000).

    Article  CAS  Google Scholar 

  6. Rodrigue, A. et al. Interplay between human DNA repair proteins at a unique double-strand break in vivo. EMBO J. 25, 222–231 (2006).

    Article  CAS  Google Scholar 

  7. Muscarella, D. E., Ellison, E. L., Ruoff, B. M. & Vogt, V. M. Characterization of I-Ppo, an intron-encoded endonuclease that mediates homing of a group I intron in the ribosomal DNA of Physarum polycephalum. Mol. Cell Biol. 10, 3386–3396 (1990).

    Article  CAS  Google Scholar 

  8. Flick, K. E., Jurica, M. S., Monnat, R. J., Jr. & Stoddard, B. L. DNA binding and cleavage by the nuclear intron-encoded homing endonuclease I-PpoI. Nature 394, 96–101 (1998).

    Article  CAS  Google Scholar 

  9. Monnat, R. J., Jr., Hackmann, A. F. & Cantrell, M. A. Generation of highly site-specific DNA double-strand breaks in human cells by the homing endonucleases I-PpoI and I-CreI. Biochem. Biophys. Res. Commun. 255, 88–93 (1999).

    Article  CAS  Google Scholar 

  10. Vigo, E. et al. CDC25A phosphatase is a target of E2F and is required for efficient E2F-induced S phase. Mol. Cell Biol. 19, 6379–6395 (1999).

    Article  CAS  Google Scholar 

  11. Berkovich, E., Lamed, Y. & Ginsberg, D. E2F and Ras synergize in transcriptionally activating p14ARF expression. Cell Cycle 2, 127–133 (2003).

    Article  CAS  Google Scholar 

  12. Bakkenist, C. J. & Kastan, M. B. DNA damage activates ATM through intermolecular autophosphorylation and dimer dissociation. Nature 421, 499–506 (2003).

    Article  CAS  Google Scholar 

  13. Kastan, M. B. & Bartek, J. Cell-cycle checkpoints and cancer. Nature 432, 316–323 (2004).

    Article  CAS  Google Scholar 

  14. Lee, J. H. & Paull, T. T. ATM activation by DNA double-strand breaks through the Mre11–Rad50–Nbs1 complex. Science 308, 551–554 (2005).

    Article  CAS  Google Scholar 

  15. Pellegrini, M. et al. Autophosphorylation at serine 1987 is dispensable for murine Atm activation in vivo. Nature 443, 222–225 (2006).

    Article  CAS  Google Scholar 

  16. Hickson, I. et al. Identification and characterization of a novel and specific inhibitor of the ataxia-telangiectasia mutated kinase ATM. Cancer Res. 64, 9152–9159 (2004).

    Article  CAS  Google Scholar 

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

    CAS  PubMed  Google Scholar 

  18. Kitagawa, R., Bakkenist, C. J., McKinnon, P. J. & Kastan, M. B. Phosphorylation of SMC1 is a critical downstream event in the ATM–NBS1–BRCA1 pathway. Genes Dev. 18, 1423–1438 (2004).

    Article  CAS  Google Scholar 

  19. Bekker-Jensen, S. et al. Spatial organization of the mammalian genome surveillance machinery in response to DNA strand breaks. J. Cell Biol. 173, 195–206 (2006).

    Article  CAS  Google Scholar 

  20. Shroff, R. et al. Distribution and dynamics of chromatin modification induced by a defined DNA double-strand break. Curr. Biol. 14, 1703–1711 (2004).

    Article  CAS  Google Scholar 

  21. Falck, J., Coates, J. & Jackson, S. P. Conserved modes of recruitment of ATM, ATR and DNA-PKcs to sites of DNA damage. Nature 434, 605–611 (2005).

    Article  CAS  Google Scholar 

  22. Young, D. B. et al. Identification of domains of Ataxia-telangiectasia Mutated required for nuclear localization and chromatin association. J. Biol. Chem. 280, 27587–27594 (2005).

    Article  CAS  Google Scholar 

  23. Collis, S. J. et al. Evasion of early cellular response mechanisms following low level radiation-induced DNA damage. J. Biol. Chem. 279, 49624–49632 (2004).

    Article  CAS  Google Scholar 

  24. Difilippantonio, S. et al. Role of Nbs1 in the activation of the Atm kinase revealed in humanized mouse models. Nature Cell Biol. 7, 675–685 (2005).

    Article  CAS  Google Scholar 

  25. Gong, F., Kwon, Y. & Smerdon, M. J. Nucleotide excision repair in chromatin and the right of entry. DNA Repair 4, 884–896 (2005).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  27. Berkovich, E. & Ginsberg, D. Ras induces elevation of E2F-1 mRNA levels. J. Biol. Chem. 276, 42851–42856 (2001).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank D. Woods and M. Reis for excellent technical assistance. This work was supported by grants from the National Institutes of Health (CA71387 and CA21765 to M.B.K. and CA48022 to R.J.M., Jr.) and by the American Lebanese Syrian Associated Charities (ALSAC) of the St. Jude Children's Research Hospital.

Author information

Authors and Affiliations

  1. Department of Oncology, St. Jude Children's Research Hospital, Memphis, 38105, TN, USA

    Elijahu Berkovich & Michael B. Kastan

  2. Departments of Pathology and of Genome Sciences, University of Washington, Box 357705, Seattle, 98195–7705, WA, USA

    Raymond J. Monnat Jr.

Authors
  1. Elijahu Berkovich
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Raymond J. Monnat Jr.
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Michael B. Kastan
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Michael B. Kastan.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary figures S1, S2, S3, S4 and S5 and Supplementary Table S1 (PDF 1729 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Berkovich, E., Monnat, R. & Kastan, M. Roles of ATM and NBS1 in chromatin structure modulation and DNA double-strand break repair. Nat Cell Biol 9, 683–690 (2007). https://doi.org/10.1038/ncb1599

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ncb1599

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
pFad - Phonifier reborn

Pfad - The Proxy pFad of © 2024 Garber Painting. All rights reserved.

Note: This service is not intended for secure transactions such as banking, social media, email, or purchasing. Use at your own risk. We assume no liability whatsoever for broken pages.


Alternative Proxies:

Alternative Proxy

pFad Proxy

pFad v3 Proxy

pFad v4 Proxy