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.

  • Article
  • Published:

53BP1 is a reader of the DNA-damage-induced H2A Lys 15 ubiquitin mark

Nature volume 499, pages 50–54 (2013)Cite this article

Subjects

Abstract

53BP1 (also called TP53BP1) is a chromatin-associated factor that promotes immunoglobulin class switching and DNA double-strand-break (DSB) repair by non-homologous end joining. To accomplish its function in DNA repair, 53BP1 accumulates at DSB sites downstream of the RNF168 ubiquitin ligase. How ubiquitin recruits 53BP1 to break sites remains unknown as its relocalization involves recognition of histone H4 Lys 20 (H4K20) methylation by its Tudor domain. Here we elucidate how vertebrate 53BP1 is recruited to the chromatin that flanks DSB sites. We show that 53BP1 recognizes mononucleosomes containing dimethylated H4K20 (H4K20me2) and H2A ubiquitinated on Lys 15 (H2AK15ub), the latter being a product of RNF168 action on chromatin. 53BP1 binds to nucleosomes minimally as a dimer using its previously characterized methyl-lysine-binding Tudor domain and a carboxy-terminal extension, termed the ubiquitination-dependent recruitment (UDR) motif, which interacts with the epitope formed by H2AK15ub and its surrounding residues on the H2A tail. 53BP1 is therefore a bivalent histone modification reader that recognizes a histone ‘code’ produced by DSB signalling.

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: Identification of the 53BP1 UDR.
Figure 2: 53BP1 binds to ubiquitinated nucleosomes.
Figure 3: 53BP1 is a bivalent reader of the H4K20me2 and H2AK15ub histone marks.
Figure 4: Determinants of H2AK15ub recognition.

Similar content being viewed by others

References

  1. Lukas, J., Lukas, C. & Bartek, J. More than just a focus: The chromatin response to DNA damage and its role in genome integrity maintenance. Nature Cell Biol. 13, 1161–1169 (2011)

    Article  CAS  Google Scholar 

  2. Jackson, S. P. & Durocher, D. Regulation of DNA damage responses by ubiquitin and SUMO. Mol. Cell 49, 795–807 (2013)

    Article  CAS  Google Scholar 

  3. Nakamura, K. et al. Genetic dissection of vertebrate 53BP1: a major role in non-homologous end joining of DNA double strand breaks. DNA Repair 5, 741–749 (2006)

    Article  CAS  Google Scholar 

  4. Bunting, S. F. et al. 53BP1 inhibits homologous recombination in Brca1-deficient cells by blocking resection of DNA breaks. Cell 141, 243–254 (2010)

    Article  CAS  Google Scholar 

  5. Ward, I. M. et al. 53BP1 is required for class switch recombination. J. Cell Biol. 165, 459–464 (2004)

    Article  CAS  Google Scholar 

  6. Manis, J. P. et al. 53BP1 links DNA damage-response pathways to immunoglobulin heavy chain class-switch recombination. Nature Immunol. 5, 481–487 (2004)

    Article  CAS  Google Scholar 

  7. Dimitrova, N., Chen, Y. C., Spector, D. L. & de Lange, T. 53BP1 promotes non-homologous end joining of telomeres by increasing chromatin mobility. Nature 456, 524–528 (2008)

    Article  ADS  CAS  Google Scholar 

  8. Bouwman, P. et al. 53BP1 loss rescues BRCA1 deficiency and is associated with triple-negative and BRCA-mutated breast cancers. Nature Struct. Mol. Biol. 17, 688–695 (2010)

    Article  CAS  Google Scholar 

  9. Bothmer, A. et al. Regulation of DNA end joining, resection, and immunoglobulin class switch recombination by 53BP1. Mol. Cell 42, 319–329 (2011)

    Article  CAS  Google Scholar 

  10. Zimmermann, M., Lottersberger, F., Buonomo, S. B., Sfeir, A. & de Lange, T. 53BP1 regulates DSB repair using Rif1 to control 5′ end resection. Science 339, 700–704 (2013)

    Article  ADS  CAS  Google Scholar 

  11. Escribano-Díaz, C. et al. A cell cycle-dependent regulatory circuit composed of 53BP1–RIF1 and BRCA1-CtIP controls DNA repair pathway choice. Mol. Cell 49, 872–883 (2013)

    Article  Google Scholar 

  12. Di Virgilio, M. et al. Rif1 prevents resection of DNA breaks and promotes immunoglobulin class switching. Science 339, 711–715 (2013)

    Article  ADS  CAS  Google Scholar 

  13. Chapman, J. R. et al. RIF1 is essential for 53BP1-dependent nonhomologous end joining and suppression of DNA double-strand break resection. Mol. Cell 49, 858–871 (2013)

    Article  CAS  Google Scholar 

  14. Botuyan, M. V. et al. Structural basis for the methylation state-specific recognition of histone H4–K20 by 53BP1 and Crb2 in DNA repair. Cell 127, 1361–1373 (2006)

    Article  CAS  Google Scholar 

  15. Huyen, Y. et al. Methylated lysine 79 of histone H3 targets 53BP1 to DNA double-strand breaks. Nature 432, 406–411 (2004)

    Article  ADS  CAS  Google Scholar 

  16. Sanders, S. L. et al. Methylation of histone H4 lysine 20 controls recruitment of Crb2 to sites of DNA damage. Cell 119, 603–614 (2004)

    Article  CAS  Google Scholar 

  17. Pei, H. et al. MMSET regulates histone H4K20 methylation and 53BP1 accumulation at DNA damage sites. Nature 470, 124–128 (2011)

    Article  ADS  CAS  Google Scholar 

  18. Acs, K. et al. The AAA-ATPase VCP/p97 promotes 53BP1 recruitment by removing L3MBTL1 from DNA double-strand breaks. Nature Struct. Mol. Biol. 18, 1345–1350 (2011)

    Article  CAS  Google Scholar 

  19. Mallette, F. A. et al. RNF8- and RNF168-dependent degradation of KDM4A/JMJD2A triggers 53BP1 recruitment to DNA damage sites. EMBO J. 31, 1865–1878 (2012)

    Article  CAS  Google Scholar 

  20. Nakamura, T. M., Moser, B. A., Du, L. L. & Russell, P. Cooperative control of Crb2 by ATM family and Cdc2 kinases is essential for the DNA damage checkpoint in fission yeast. Mol. Cell. Biol. 25, 10721–10730 (2005)

    Article  CAS  Google Scholar 

  21. Zgheib, O., Pataky, K., Brugger, J. & Halazonetis, T. D. An oligomerized 53BP1 tudor domain suffices for recognition of DNA double-strand breaks. Mol. Cell. Biol. 29, 1050–1058 (2009)

    Article  CAS  Google Scholar 

  22. Pryde, F. et al. 53BP1 exchanges slowly at the sites of DNA damage and appears to require RNA for its association with chromatin. J. Cell Sci. 118, 2043–2055 (2005)

    Article  CAS  Google Scholar 

  23. Gudjonsson, T. et al. TRIP12 and UBR5 suppress spreading of chromatin ubiquitylation at damaged chromosomes. Cell 150, 697–709 (2012)

    Article  CAS  Google Scholar 

  24. Stewart, G. S. et al. The RIDDLE syndrome protein mediates a ubiquitin-dependent signaling cascade at sites of DNA damage. Cell 136, 420–434 (2009)

    Article  CAS  Google Scholar 

  25. Panier, S. et al. Tandem protein interaction modules organize the ubiquitin-dependent response to DNA double-strand breaks. Mol. Cell 47, 383–395 (2012)

    Article  CAS  Google Scholar 

  26. Mattiroli, F. et al. RNF168 ubiquitinates K13–15 on H2A/H2AX to drive DNA damage signaling. Cell 150, 1182–1195 (2012)

    Article  CAS  Google Scholar 

  27. Gatti, M. et al. A novel ubiquitin mark at the N-terminal tail of histone H2As targeted by RNF168 ubiquitin ligase. Cell Cycle 11, 2538–2544 (2012)

    Article  CAS  Google Scholar 

  28. Wang, H. et al. Role of histone H2A ubiquitination in Polycomb silencing. Nature 431, 873–878 (2004)

    Article  ADS  CAS  Google Scholar 

  29. Simon, M. D. et al. The site-specific installation of methyl-lysine analogs into recombinant histones. Cell 128, 1003–1012 (2007)

    Article  CAS  Google Scholar 

  30. Dikic, I., Wakatsuki, S. & Walters, K. J. Ubiquitin-binding domains—from structures to functions. Nature Rev. Mol. Cell Biol. 10, 659–671 (2009)

    Article  CAS  Google Scholar 

  31. Chen, J., Ai, Y., Wang, J., Haracska, L. & Zhuang, Z. Chemically ubiquitylated PCNA as a probe for eukaryotic translesion DNA synthesis. Nature Chem. Biol. 6, 270–272 (2010)

    Article  CAS  Google Scholar 

  32. Wu, L. et al. ASH2L regulates ubiquitylation signaling to MLL: trans-regulation of H3 K4 methylation in higher eukaryotes. Mol. Cell 49, 1108–1120 (2013)

    Article  CAS  Google Scholar 

  33. Poulsen, M., Lukas, C., Lukas, J., Bekker-Jensen, S. & Mailand, N. Human RNF169 is a negative regulator of the ubiquitin-dependent response to DNA double-strand breaks. J. Cell Biol. 197, 189–199 (2012)

    Article  CAS  Google Scholar 

  34. Juang, Y. C. et al. OTUB1 co-opts Lys48-linked ubiquitin recognition to suppress E2 enzyme function. Mol. Cell 45, 384–397 (2012)

    Article  CAS  Google Scholar 

  35. Morita, S., Kojima, T. & Kitamura, T. Plat-E: an efficient and stable system for transient packaging of retroviruses. Gene Ther. 7, 1063–1066 (2000)

    Article  CAS  Google Scholar 

  36. Dyer, P. N. et al. Reconstitution of nucleosome core particles from recombinant histones and DNA. Methods Enzymol. 375, 23–44 (2004)

    Article  CAS  Google Scholar 

  37. Luger, K., Mader, A. W., Richmond, R. K., Sargent, D. F. & Richmond, T. J. Crystal structure of the nucleosome core particle at 2.8 Å resolution. Nature 389, 251–260 (1997)

    Article  ADS  CAS  Google Scholar 

  38. Nakamura, T. M., Du, L. L., Redon, C. & Russell, P. Histone H2A phosphorylation controls Crb2 recruitment at DNA breaks, maintains checkpoint arrest, and influences DNA repair in fission yeast. Mol. Cell. Biol. 24, 6215–6230 (2004)

    Article  CAS  Google Scholar 

  39. Bentley, M. L. et al. Recognition of UbcH5c and the nucleosome by the Bmi1/Ring1b ubiquitin ligase complex. EMBO J. 30, 3285–3297 (2011)

    Article  CAS  Google Scholar 

  40. Wang, A. C., Grzesiek, S., Tschudin, R., Lodi, P. J. & Bax, A. Sequential backbone assignment of isotopically enriched proteins in D2O by deuterium-decoupled HA(CA)N and HA(CACO)N. J. Biomol. NMR 5, 376–382 (1995)

    CAS  PubMed  Google Scholar 

Download references

Acknowledgements

We are grateful to R. Szilard, J. Côté and S. Panier for critically reading the manuscript; to L.-L. Du, A. Nussenzweig, Y. Tong and C. Arrowsmith for plasmids; to M. Cook and A. Rosebrock for centrifugal elutriation; and to B. Sauriol for help with FRAP data analysis. A.F.-T. and A.O. both receive post-doctoral fellowships from the CIHR; C.E.-D. is an Ontario Post-doctoral Fellow; J.K.-L. receives a post-doctoral fellowship from the Leukemia and Lymphoma Society; and C.C.Y.L. and M.-C.L. are post-doctoral fellows of the Canadian Breast Cancer Foundation (Ontario Division). D.D. is the Thomas Kierans Chair in Mechanisms of Cancer Development and a Canada Research Chair (Tier 1) in the Molecular Mechanisms of Genome Integrity. Work in the D.D. laboratory was supported by CIHR grant MOP84297 and grant GL2-01-010 from the Ontario Research Fund.

Author information

Authors and Affiliations

  1. Samuel Lunenfeld Research Institute, Mount Sinai Hospital, 600 University Avenue, Toronto, M5G 1X5, Ontario, Canada

    Amélie Fradet-Turcotte, Marella D. Canny, Cristina Escribano-Díaz, Alexandre Orthwein, Charles C. Y. Leung, Hao Huang, Marie-Claude Landry, Sylvie M. Noordermeer, Frank Sicheri & Daniel Durocher

  2. Department of Molecular Genetics, University of Toronto, M5S 3E1, Ontario, Canada

    Julianne Kitevski-LeBlanc, Frank Sicheri & Daniel Durocher

  3. Department of Biochemistry, University of Toronto, M5S 3E1, Ontario, Canada

    Julianne Kitevski-LeBlanc & Frank Sicheri

  4. Department of Chemistry, University of Toronto, M5S 3E1, Ontario, Canada

    Julianne Kitevski-LeBlanc

Authors
  1. Amélie Fradet-Turcotte
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Marella D. Canny
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Cristina Escribano-Díaz
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Alexandre Orthwein
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Charles C. Y. Leung
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Hao Huang
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Marie-Claude Landry
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Julianne Kitevski-LeBlanc
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Sylvie M. Noordermeer
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Frank Sicheri
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Daniel Durocher
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

A.F.-T. initiated the project, carried out most of the cell biological experiments, found the interaction between ubiquitinated H2A and 53BP1, and contributed to the experimental design and data interpretation. M.D.C. produced the recombinant nucleosomes. A.F.-T. and M.D.C. and carried out the recombinant nucleosome pull-down studies. C.E.-D. examined RIF1 focus formation, the role of the UDR in homologous recombination and helped with some immunofluorescence experiments. A.O. carried out the class switching experiments and the DT40 work. C.C.Y.L. helped with protein binding studies and generated the nucleosome model shown in Supplementary Fig. 12. M.-C.L. helped with the FRAP experiments. J.K.-L. carried out chemical ubiquitination and mass spectrometry to verify histone methylation. H.H. carried out the NMR experiments. S.M.N. carried out mass spectrometry to verify H4K20 methylation. F.S. supervised H.H. D.D. supervised the project and wrote the manuscript with input from the other authors.

Corresponding author

Correspondence to Daniel Durocher.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

This file contains Supplementary Figures 1-14, Supplementary Text and Data and additional references. (PDF 10429 kb)

PowerPoint slides

Source data

Rights and permissions

Reprints and permissions

About this article

Cite this article

Fradet-Turcotte, A., Canny, M., Escribano-Díaz, C. et al. 53BP1 is a reader of the DNA-damage-induced H2A Lys 15 ubiquitin mark. Nature 499, 50–54 (2013). https://doi.org/10.1038/nature12318

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature12318

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