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The SUMO modification pathway is involved in the BRCA1 response to genotoxic stress

Nature volume 462, pages 886–890 (2009)Cite this article

Abstract

Mutations in BRCA1 are associated with a high risk of breast and ovarian cancer. BRCA1 participates in the DNA damage response and acts as a ubiquitin ligase. However, its regulation remains poorly understood. Here we report that BRCA1 is modified by small ubiquitin-like modifier (SUMO) in response to genotoxic stress, and co-localizes at sites of DNA damage with SUMO1, SUMO2/3 and the SUMO-conjugating enzyme Ubc9. PIAS SUMO E3 ligases co-localize with and modulate SUMO modification of BRCA1, and are required for BRCA1 ubiquitin ligase activity in cells. In vitro SUMO modification of the BRCA1/BARD1 heterodimer greatly increases its ligase activity, identifying it as a SUMO-regulated ubiquitin ligase (SRUbL). Further, PIAS SUMO ligases are required for complete accumulation of double-stranded DNA (dsDNA) damage-repair proteins subsequent to RNF8 accrual, and for proficient double-strand break repair. These data demonstrate that the SUMOylation pathway plays a significant role in mammalian DNA damage response.

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Figure 1: The SUMO-conjugation machinery locates to sites of DNA damage, and BRCA1 is modified by SUMO after genotoxic stress.
Figure 2: PIAS E3 SUMO ligases modulate BRCA1 SUMOylation.
Figure 3: PIAS E3 SUMO ligases influence accumulation of DNA damage-repair protein and are required for dsDNA break repair.
Figure 4: The SUMO pathway regulates BRCA1 ubiquitin ligase activity.

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References

  1. Lorick, K. L. et al. RING fingers mediate ubiquitin-conjugating enzyme (E2)-dependent ubiquitination. Proc. Natl Acad. Sci. USA 96, 11364–11369 (1999)

    Article  ADS  CAS  Google Scholar 

  2. Hashizume, R. et al. The RING heterodimer BRCA1-BARD1 is a ubiquitin ligase inactivated by a breast cancer-derived mutation. J. Biol. Chem. 276, 14537–14540 (2001)

    Article  CAS  Google Scholar 

  3. Brzovic, P. S. et al. Binding and recognition in the assembly of an active BRCA1/BARD1 ubiquitin-ligase complex. Proc. Natl Acad. Sci. USA 100, 5646–5651 (2003)

    Article  ADS  CAS  Google Scholar 

  4. Morris, J. R. et al. Genetic analysis of BRCA1 ubiquitin ligase activity and its relationship to breast cancer susceptibility. Hum. Mol. Genet. 15, 599–606 (2006)

    Article  CAS  Google Scholar 

  5. Morris, J. R. & Solomon, E. BRCA1: BARD1 induces the formation of conjugated ubiquitin structures, dependent on K6 of ubiquitin, in cells during DNA replication and repair. Hum. Mol. Genet. 13, 807–817 (2004)

    Article  CAS  Google Scholar 

  6. Morris, J. R. & Solomon, E. in The Role of Genetics in Breast and Reproductive Cancers (ed. Welcsh, P. L.) 75–92 (Springer Science+Business Media and Humana Press, 2009)

    Google Scholar 

  7. Reid, L. J. et al. E3 ligase activity of BRCA1 is not essential for mammalian cell viability or homology-directed repair of double-strand DNA breaks. Proc. Natl Acad. Sci. USA 105, 20876–20881 (2008)

    Article  ADS  CAS  Google Scholar 

  8. Polanowska, J., Martin, J. S., Garcia-Muse, T., Petalcorin, M. I. & Boulton, S. J. A conserved pathway to activate BRCA1-dependent ubiquitylation at DNA damage sites. EMBO J. 25, 2178–2188 (2006)

    Article  CAS  Google Scholar 

  9. Zhao, G. Y. et al. A critical role for the ubiquitin-conjugating enzyme Ubc13 in initiating homologous recombination. Mol. Cell 25, 663–675 (2007)

    Article  CAS  Google Scholar 

  10. Doil, C. et al. RNF168 binds and amplifies ubiquitin conjugates on damaged chromosomes to allow accumulation of repair proteins. Cell 136, 435–446 (2009)

    Article  CAS  Google Scholar 

  11. Huen, M. S. et al. RNF8 transduces the DNA-damage signal via histone ubiquitylation and checkpoint protein assembly. Cell 131, 901–914 (2007)

    Article  CAS  Google Scholar 

  12. Kolas, N. K. et al. Orchestration of the DNA-damage response by the RNF8 ubiquitin ligase. Science 318, 1637–1640 (2007)

    Article  ADS  CAS  Google Scholar 

  13. Mailand, N. et al. RNF8 ubiquitylates histones at DNA double-strand breaks and promotes assembly of repair proteins. Cell 131, 887–900 (2007)

    Article  CAS  Google Scholar 

  14. Wang, B. & Elledge, S. J. Ubc13/Rnf8 ubiquitin ligases control foci formation of the Rap80/Abraxas/Brca1/Brcc36 complex in response to DNA damage. Proc. Natl Acad. Sci. USA 104, 20759–20763 (2007)

    Article  ADS  CAS  Google Scholar 

  15. Kim, H., Huang, J. & Chen, J. CCDC98 is a BRCA1-BRCT domain-binding protein involved in the DNA damage response. Nature Struct. Mol. Biol. 14, 710–715 (2007)

    Article  CAS  Google Scholar 

  16. Kim, H., Chen, J. & Yu, X. Ubiquitin-binding protein RAP80 mediates BRCA1-dependent DNA damage response. Science 316, 1202–1205 (2007)

    Article  ADS  CAS  Google Scholar 

  17. Liu, Z., Wu, J. & Yu, X. CCDC98 targets BRCA1 to DNA damage sites. Nature Struct. Mol. Biol. 14, 716–720 (2007)

    Article  CAS  Google Scholar 

  18. Sobhian, B. et al. RAP80 targets BRCA1 to specific ubiquitin structures at DNA damage sites. Science 316, 1198–1202 (2007)

    Article  ADS  CAS  Google Scholar 

  19. Wang, B. et al. Abraxas and RAP80 form a BRCA1 protein complex required for the DNA damage response. Science 316, 1194–1198 (2007)

    Article  ADS  CAS  Google Scholar 

  20. Yan, J. et al. The ubiquitin-interacting motif containing protein RAP80 interacts with BRCA1 and functions in DNA damage repair response. Cancer Res. 67, 6647–6656 (2007)

    Article  CAS  Google Scholar 

  21. 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 

  22. Hay, R. T. SUMO: a history of modification. Mol. Cell 18, 1–12 (2005)

    Article  CAS  Google Scholar 

  23. Mo, Y. Y., Yu, Y., Ee, P. L. & Beck, W. T. Overexpression of a dominant-negative mutant Ubc9 is associated with increased sensitivity to anticancer drugs. Cancer Res. 64, 2793–2798 (2004)

    Article  CAS  Google Scholar 

  24. Zhao, X. & Blobel, G. A. SUMO ligase is part of a nuclear multiprotein complex that affects DNA repair and chromosomal organization. Proc. Natl Acad. Sci. USA 102, 4777–4782 (2005)

    Article  ADS  CAS  Google Scholar 

  25. Potts, P. R. & Yu, H. Human MMS21/NSE2 is a SUMO ligase required for DNA repair. Mol. Cell. Biol. 25, 7021–7032 (2005)

    Article  CAS  Google Scholar 

  26. Mabb, A. M., Wuerzberger-Davis, S. M. & Miyamoto, S. PIASy mediates NEMO sumoylation and NF-κB activation in response to genotoxic stress. Nature Cell Biol. 8, 986–993 (2006)

    Article  CAS  Google Scholar 

  27. Ishiai, M. et al. DNA cross-link repair protein SNM1A interacts with PIAS1 in nuclear focus formation. Mol. Cell. Biol. 24, 10733–10741 (2004)

    Article  CAS  Google Scholar 

  28. Boulton, S. J. et al. BRCA1/BARD1 orthologs required for DNA repair in Caenorhabditis elegans . Curr. Biol. 14, 33–39 (2004)

    Article  CAS  Google Scholar 

  29. Park, M. A., Seok, Y. J., Jeong, G. & Lee, J. S. SUMO1 negatively regulates BRCA1-mediated transcription, via modulation of promoter occupancy. Nucleic Acids Res. 36, 263–283 (2008)

    Article  CAS  Google Scholar 

  30. Peter, M. & Ameer-Beg, S. M. Imaging molecular interactions by multiphoton FLIM. Biol. Cell 96, 231–236 (2004)

    Article  CAS  Google Scholar 

  31. Peter, M. et al. Multiphoton-FLIM quantification of the EGFP-mRFP1 FRET pair for localization of membrane receptor-kinase interactions. Biophys. J. 88, 1224–1237 (2005)

    Article  ADS  CAS  Google Scholar 

  32. Ng, T. et al. Imaging protein kinase Cα activation in cells. Science 283, 2085–2089 (1999)

    Article  ADS  CAS  Google Scholar 

  33. Ganesan, S., Ameer-Beg, S. M., Ng, T. T., Vojnovic, B. & Wouters, F. S. A dark yellow fluorescent protein (YFP)-based resonance energy-accepting chromoprotein (REACh) for Forster resonance energy transfer with GFP. Proc. Natl Acad. Sci. USA 103, 4089–4094 (2006)

    Article  ADS  CAS  Google Scholar 

  34. Dadke, S. et al. Regulation of protein tyrosine phosphatase 1B by sumoylation. Nature Cell Biol. 9, 80–85 (2007)

    Article  CAS  Google Scholar 

  35. Bossis, G. & Melchior, F. Regulation of SUMOylation by reversible oxidation of SUMO conjugating enzymes. Mol. Cell 21, 349–357 (2006)

    Article  CAS  Google Scholar 

  36. Schmidt, D. & Muller, S. PIAS/SUMO: new partners in transcriptional regulation. Cell. Mol. Life Sci. 60, 2561–2574 (2003)

    Article  CAS  Google Scholar 

  37. Munarriz, E. et al. PIAS-1 is a checkpoint regulator which affects exit from G1 and G2 by sumoylation of p73. Mol. Cell. Biol. 24, 10593–10610 (2004)

    Article  CAS  Google Scholar 

  38. Kim, H. & Chen, J. New players in the BRCA1-mediated DNA damage responsive pathway. Mol. Cells 25, 457–461 (2008)

    CAS  PubMed  PubMed Central  Google Scholar 

  39. Nishikawa, H., Ooka, S., Sato, K., Arima, K., Okamoto, J., Klevit, R. E., Fukuda, M. & Ohta, T. Mass spectrometric and mutational analyses reveal Lys-6-linked polyubiquitin chains catalyzed by BRCA1-BARD1 ubiquitin ligase. J Biol Chem (2003)

  40. Wu-Baer, F., Lagrazon, K., Yuan, W. & Baer, R. The BRCA1/BARD1 heterodimer assembles polyubiquitin chains through an unconventional linkage involving lysine residue K6 of ubiquitin. J. Biol. Chem. 278, 34743–34746 (2003)

    Article  CAS  Google Scholar 

  41. Mallery, D. L., Vandenberg, C. J. & Hiom, K. Activation of the E3 ligase function of the BRCA1/BARD1 complex by polyubiquitin chains. EMBO J. 21, 6755–6762 (2002)

    Article  CAS  Google Scholar 

  42. Christensen, D. E., Brzovic, P. S. & Klevit, R. E. E2-BRCA1 RING interactions dictate synthesis of mono- or specific polyubiquitin chain linkages. Nature Struct. Mol. Biol. 14, 941–948 (2007)

    Article  CAS  Google Scholar 

  43. Manke, I. A., Lowery, D. M., Nguyen, A. & Yaffe, M. B. BRCT repeats as phosphopeptide-binding modules involved in protein targeting. Science 302, 636–639 (2003)

    Article  ADS  CAS  Google Scholar 

  44. Jaffray, E. G. & Hay, R. T. Detection of modification by ubiquitin-like proteins. Methods 38, 35–38 (2006)

    Article  CAS  Google Scholar 

  45. Prag, S. et al. Activated ezrin promotes cell migration through recruitment of the GEF Dbl to lipid rafts and preferential downstream activation of Cdc42. Mol. Biol. Cell 18, 2935–2948 (2007)

    Article  CAS  Google Scholar 

  46. Barber, P. R., Ameer-Beg, S. M., Gilbey, J. D., Edens, R. J., Ezike, I. & Vojnovic, B. Global and pixel kinetic data analysis for FRET detection by multi-photon time-domain FLIM. Proc. SPIE 5700, 171–181 (2005)

    Article  ADS  CAS  Google Scholar 

  47. Bennardo, N., Cheng, A., Huang, N. & Stark, J. M. Alternative-NHEJ is a mechanistically distinct pathway of mammalian chromosome break repair. PLoS Genet. 4, e1000110 (2008)

    Article  Google Scholar 

  48. Boutell, C., Orr, A. & Everett, R. D. PML residue lysine 160 is required for the degradation of PML induced by herpes simplex virus type 1 regulatory protein ICP0. J. Virol. 77, 8686–8694 (2003)

    Article  Google Scholar 

  49. Boutell, C., Sadis, S. & Everett, R. D. Herpes simplex virus type 1 immediate-early protein ICP0 and its isolated RING finger domain act as ubiquitin E3 ligases in vitro . J. Virol. 76, 841–850 (2002)

    Article  CAS  Google Scholar 

  50. Bliss, C. I. The toxicity of poisons applied jointly. Ann. Appl. Biol. 26, 585–615 (1939)

    Article  CAS  Google Scholar 

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Acknowledgements

We are grateful to V. De Laurenzi for the PIAS1 expression constructs, to R. Hay for His-SUMO1 and His-SUMO2 cells, to D. Durocher for anti-RNF168 antibody, to G. Stewart for discussions and S. Jackson for sharing results before publication. The work was supported by grants from Breast Cancer Campaign (to J.R.M., A.A., #SF06, and L.B., #06NovPHD13Morris), Cancer Research UK (to R.D., #C8820/A9494), the Medical Research Council (to E.S. and D.W., #6900577, and C.B.), the Richard Dimbleby Cancer Fund to King’s College London (to M.K. and T.N.) and Breakthrough Breast Cancer (to T.K). Multiphoton FLIM systems and acquisition/analysis software were built by S. Ameer-Beg, P. Barber and B. Vojnovic, with support from MRC Co-operative Group Grant G0100152 #56891 and UK Research Councils Basic Technology Research Programme Grant GR/R87901/01.

Author Contributions J.R.M. conceived and designed the study, generated reagents, performed experiments and wrote the paper. C.B. performed in vitro assays, confirmed and developed the initial concept, and generated reagents. M.K. optimised and performed FLIM measurements and analysis. R.D. and D.W. performed experiments and generated reagents. L.B. performed co-localisation observations, A.A. and L.P. generated reagents and Y.G. generated reagents and participated in discussions. T.K. undertook FLIM measurements. T.N. provided expertise and input into the design of the FLIM experiments, and E.S. provided advice and mentoring to J.R.M.

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Author notes

  1. Joanna R. Morris, Chris Boutell and Melanie Keppler: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Medical and Molecular Genetics, King’s College London, Guy’s Medical School Campus, London SE1 9RT, UK

    Joanna R. Morris, Ruth Densham, Daniel Weekes, Amin Alamshah, Laura Butler, Laurent Pangon & Ellen Solomon

  2. MRC Virology Unit, Church Street, Glasgow G11 5JR, Scotland, UK ,

    Chris Boutell

  3. Richard Dimbleby Department of Cancer Research, Randall Division of Cell and Molecular Biophysics, King’s College London, Guy’s Medical School Campus, London SE1 1UL, UK

    Melanie Keppler, Tai Kiuchi & Tony Ng

  4. The Wellcome Trust/Cancer Research UK Gurdon Institute and Department of Zoology, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK,

    Yaron Galanty

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Morris, J., Boutell, C., Keppler, M. et al. The SUMO modification pathway is involved in the BRCA1 response to genotoxic stress. Nature 462, 886–890 (2009). https://doi.org/10.1038/nature08593

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