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Linear ubiquitination prevents inflammation and regulates immune signalling

Nature volume 471, pages 591–596 (2011)Cite this article

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Abstract

Members of the tumour necrosis factor (TNF) receptor superfamily have important functions in immunity and inflammation. Recently linear ubiquitin chains assembled by a complex containing HOIL-1 and HOIP (also known as RBCK1 and RNF31, respectively) were implicated in TNF signalling, yet their relevance in vivo remained uncertain. Here we identify SHARPIN as a third component of the linear ubiquitin chain assembly complex, recruited to the CD40 and TNF receptor signalling complexes together with its other constituents, HOIL-1 and HOIP. Mass spectrometry of TNF signalling complexes revealed RIP1 (also known as RIPK1) and NEMO (also known as IKKγ or IKBKG) to be linearly ubiquitinated. Mutation of the Sharpin gene (Sharpincpdm/cpdm) causes chronic proliferative dermatitis (cpdm) characterized by inflammatory skin lesions and defective lymphoid organogenesis. Gene induction by TNF, CD40 ligand and interleukin-1β was attenuated in cpdm-derived cells which were rendered sensitive to TNF-induced death. Importantly, Tnf gene deficiency prevented skin lesions in cpdm mice. We conclude that by enabling linear ubiquitination in the TNF receptor signalling complex, SHARPIN interferes with TNF-induced cell death and, thereby, prevents inflammation. Our results provide evidence for the relevance of linear ubiquitination in vivo in preventing inflammation and regulating immune signalling.

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Figure 1: SHARPIN is recruited to the native CD40- and TNF-RSCs.
Figure 2: SHARPIN forms a tripartite complex with HOIP and HOIL-1 that is capable of forming linear ubiquitin chains.
Figure 3: NEMO and RIP1 are linearly ubiquitinated in the native TNF-RSC.
Figure 4: SHARPIN is required for full TNF-, CD40L- and IL-1β-induced activation of NF-κB and JNK and to prevent TNF-induced cell death.
Figure 5: Genetic ablation of TNF rescues the skin phenotype and reduces inflammation in cpdm mice but does not revert the immunological distortion.

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References

  1. Ware, C. F. The TNF superfamily. Cytokine Growth Factor Rev. 14, 181–184 (2003)

    Article  CAS  Google Scholar 

  2. Bianchi, K. & Meier, P. A tangled web of ubiquitin chains: breaking news in TNF-R1 signaling. Mol. Cell 36, 736–742 (2009)

    Article  CAS  Google Scholar 

  3. Silke, J. & Brink, R. Regulation of TNFRSF and innate immune signalling complexes by TRAFs and cIAPs. Cell Death Differ. 17, 35–45 (2010)

    Article  CAS  Google Scholar 

  4. Wertz, I. E. & Dixit, V. M. Ubiquitin-mediated regulation of TNFR1 signaling. Cytokine Growth Factor Rev. 19, 313–324 (2008)

    Article  CAS  Google Scholar 

  5. Haas, T. L. et al. Recruitment of the linear ubiquitin chain assembly complex stabilizes the TNF-R1 signaling complex and is required for TNF-mediated gene induction. Mol. Cell 36, 831–844 (2009)

    Article  CAS  Google Scholar 

  6. Tokunaga, F. et al. Involvement of linear polyubiquitylation of NEMO in NF-κB activation. Nature Cell Biol. 11, 123–132 (2009)

    Article  CAS  Google Scholar 

  7. Rahighi, S. et al. Specific recognition of linear ubiquitin chains by NEMO is important for NF-κB activation. Cell 136, 1098–1109 (2009)

    Article  CAS  Google Scholar 

  8. Lo, Y. C. et al. Structural basis for recognition of diubiquitins by NEMO. Mol. Cell 33, 602–615 (2009)

    Article  CAS  Google Scholar 

  9. Komander, D. et al. Molecular discrimination of structurally equivalent Lys 63-linked and linear polyubiquitin chains. EMBO Rep. 10, 466–473 (2009)

    Article  CAS  Google Scholar 

  10. Xu, M., Skaug, B., Zeng, W. & Chen, Z. J. A ubiquitin replacement strategy in human cells reveals distinct mechanisms of IKK activation by TNFα and IL-1β. Mol. Cell 36, 302–314 (2009)

    Article  CAS  Google Scholar 

  11. Cossu, F. et al. Structural basis for bivalent Smac-mimetics recognition in the IAP protein family. J. Mol. Biol. 392, 630–644 (2009)

    Article  CAS  Google Scholar 

  12. Seymour, R. E. et al. Spontaneous mutations in the mouse Sharpin gene result in multiorgan inflammation, immune system dysregulation and dermatitis. Genes Immun. 8, 416–421 (2007)

    Article  CAS  Google Scholar 

  13. HogenEsch, H. et al. A spontaneous mutation characterized by chronic proliferative dermatitis in C57BL mice. Am. J. Pathol. 143, 972–982 (1993)

    CAS  PubMed  PubMed Central  Google Scholar 

  14. HogenEsch, H., Janke, S., Boggess, D. & Sundberg, J. P. Absence of Peyer’s patches and abnormal lymphoid architecture in chronic proliferative dermatitis (cpdm/cpdm) mice. J. Immunol. 162, 3890–3896 (1999)

    CAS  PubMed  Google Scholar 

  15. HogenEsch, H. et al. Increased expression of type 2 cytokines in chronic proliferative dermatitis (cpdm) mutant mice and resolution of inflammation following treatment with IL-12. Eur. J. Immunol. 31, 734–742 (2001)

    Article  CAS  Google Scholar 

  16. Tay, S. et al. Single-cell NF-κB dynamics reveal digital activation and analogue information processing. Nature 466, 267–271 (2010)

    Article  ADS  CAS  Google Scholar 

  17. Senftleben, U. et al. Activation by IKKα of a second, evolutionary conserved, NF-κB signaling pathway. Science 293, 1495–1499 (2001)

    Article  ADS  CAS  Google Scholar 

  18. Gijbels, M. J., HogenEsch, H., Bruijnzeel, P. L., Elliott, G. R. & Zurcher, C. Maintenance of donor phenotype after full-thickness skin transplantation from mice with chronic proliferative dermatitis (cpdm/cpdm) to C57BL/Ka and nude mice and vice versa. J. Invest. Dermatol. 105, 769–773 (1995)

    Article  CAS  Google Scholar 

  19. Liang, Y., Seymour, R. E. & Sundberg, J. P. Inhibition of NF-κB signaling retards eosinophilic dermatitis in SHARPIN-deficient mice. J. Invest. Dermatol. 131, 141–149 (2011)

    Article  CAS  Google Scholar 

  20. Martin, M. U. & Wesche, H. Summary and comparison of the signaling mechanisms of the Toll/interleukin-1 receptor family. Biochim. Biophys. Acta 1592, 265–280 (2002)

    Article  CAS  Google Scholar 

  21. Arend, W. P., Palmer, G. & Gabay, C. IL-1, IL-18, and IL-33 families of cytokines. Immunol. Rev. 223, 20–38 (2008)

    Article  CAS  Google Scholar 

  22. Huang, H. et al. K33-linked polyubiquitination of T cell receptor-ζ regulates proteolysis-independent T cell signaling. Immunity 33, 60–70 (2010)

    Article  CAS  Google Scholar 

  23. Matsumoto, M. L. et al. K11-linked polyubiquitination in cell cycle control revealed by a K11 linkage-specific antibody. Mol. Cell 39, 477–484 (2010)

    Article  CAS  Google Scholar 

  24. Arimoto, K. et al. Polyubiquitin conjugation to NEMO by triparite motif protein 23 (TRIM23) is critical in antiviral defense. Proc. Natl Acad. Sci. USA 107, 15856–15861 (2010)

    Article  ADS  CAS  Google Scholar 

  25. Ikeda, F., Crosetto, N. & Dikic, I. What determines the specificity and outcomes of ubiquitin signaling? Cell 143, 677–681 (2010)

    Article  CAS  Google Scholar 

  26. Liu, S. & Chen, Z. J. Expanding role of ubiquitination in NF-κB signaling. Cell Res. 21, 6–21 (2011)

    Article  Google Scholar 

  27. Laplantine, E. et al. NEMO specifically recognizes K63-linked poly-ubiquitin chains through a new bipartite ubiquitin-binding domain. EMBO J. 28, 2885–2895 (2009)

    Article  CAS  Google Scholar 

  28. Dynek, J. N. et al. c-IAP1 and UbcH5 promote K11-linked polyubiquitination of RIP1 in TNF signalling. EMBO J. 29, 4198–4209 (2010)

    Article  CAS  Google Scholar 

  29. Taylor, P. C. & Feldmann, M. Anti-TNF biologic agents: still the therapy of choice for rheumatoid arthritis. Nature Rev. Rheumatol. 5, 578–582 (2009)

    Article  CAS  Google Scholar 

  30. Sacre, S. M., Andreakos, E., Taylor, P., Feldmann, M. & Foxwell, B. M. Molecular therapeutic targets in rheumatoid arthritis. Expert Rev. Mol. Med. 7, 1–20 (2005)

    Article  Google Scholar 

  31. Mantovani, A., Allavena, P., Sica, A. & Balkwill, F. Cancer-related inflammation. Nature 454, 436–444 (2008)

    Article  ADS  CAS  Google Scholar 

  32. Pikarsky, E. et al. NF-κB functions as a tumour promoter in inflammation-associated cancer. Nature 431, 461–466 (2004)

    Article  ADS  CAS  Google Scholar 

  33. Grivennikov, S. I., Greten, F. R. & Karin, M. Immunity, inflammation, and cancer. Cell 140, 883–899 (2010)

    Article  CAS  Google Scholar 

  34. Silke, J. et al. Determination of cell survival by RING-mediated regulation of inhibitor of apoptosis (IAP) protein abundance. Proc. Natl Acad. Sci. USA 102, 16182–16187 (2005)

    Article  ADS  CAS  Google Scholar 

  35. Körner, H. et al. Distinct roles for lymphotoxin-α and tumor necrosis factor in organogenesis and spatial organization of lymphoid tissue. Eur. J. Immunol. 27, 2600–2609 (1997)

    Article  Google Scholar 

  36. Diessenbacher, P. et al. NF-κB inhibition reveals differential mechanisms of TNF versus TRAIL-induced apoptosis upstream or at the level of caspase-8 activation independent of cIAP2. J. Invest. Dermatol. 128, 1134–1147 (2008)

    Article  CAS  Google Scholar 

  37. Moolenbeek, C. & Ruitenberg, E. J. The ‘Swiss roll’: a simple technique for histological studies of the rodent intestine. Lab. Anim. 15, 57–59 (1981)

    Article  CAS  Google Scholar 

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Acknowledgements

We thank M. Miasari for developing the cIAP2 antibody, M. Leverkus for advice on skin histology, J. Lokan for advice on liver histology, C. Rappl, N. Barboza, S. Kupka, D. Heinze, J. Zipprich, J. Corbin, S. Wiegelmann and A. Bankovacki for excellent technical assistance, M. Bolognesi and P. Seneci for SM-164, H. Koerner for Tnf−/− mice, and all past and present members of the Walczak and Silke laboratories for continuous stimulating scientific discussions and support. Research in the Walczak lab is supported by grants from Cancer Research UK, AICR, BBSRC (ERASysBio PLUS), Ovarian Cancer Action and the EU Marie Curie Research Training Network ApopTRAIN. Work in the Silke lab is supported by NHMRC grants 541901, 541902 and 602516. Work in the Purcell lab is supported by the NHMRC (Senior Research Fellowships for A.W.P. and a C. J. Martin Overseas Biomedical Fellowship to A.I.W.), and by grants from the NHMRC and ARC. U.W. is funded by HGF/SBCancer. E.R. and T.L.H. are ApopTRAIN fellows and U.N. is supported by the Schweizer Nationalfonds (SNF).

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Author notes

  1. Björn Gerlach, Stefanie M. Cordier, Anna C. Schmukle, Christoph H. Emmerich, Eva Rieser, Tobias L. Haas and Andrew I. Webb: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Medicine, Tumour Immunology Unit, Imperial College London, W12 0NN London, UK,

    Björn Gerlach, Stefanie M. Cordier, Anna C. Schmukle, Christoph H. Emmerich, Eva Rieser & Henning Walczak

  2. Division of Apoptosis Regulation, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany,

    Björn Gerlach, Christoph H. Emmerich, Tobias L. Haas & Henning Walczak

  3. Department of Experimental Oncology, Mediterranean Institute of Oncology, 95029 Viagrande, Italy,

    Tobias L. Haas

  4. Department of Biochemistry, Bio21 Melbourne University, Melbourne, VIC 3010, Australia,

    Andrew I. Webb & Anthony W. Purcell

  5. Department of Biochemistry, La Trobe University, Melbourne, VIC 3086, Australia,

    James A. Rickard, Holly Anderton, Wendy W.-L. Wong, Ueli Nachbur, Lahiru Gangoda & John Silke

  6. Protein Analysis Core Facility, German Cancer Research Center (DKFZ), 69120 Heidelberg, Germany,

    Uwe Warnken

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Contributions

T.L.H. and H.W. conceived the moTAP procedure. B.G., E.R., H.W. and U.W., and T.L.H. and U.W. determined the composition of the CD40- and TNF-RSC, respectively. B.G. performed all other experiments involving CD40L. A.C.S. and C.H.E. cloned and purified all recombinant proteins, determined the molecular interaction between the LUBAC components and performed the in vitro ubiquitination assays. B.G., S.M.C., A.C.S., C.H.E. and E.R. provided moTAP-purified TNF-RSCs for 2D-MRM analysis which was conceived by A.I.W. and H.W., performed by A.I.W., and analysed by A.I.W. and A.W.P.; J.S. and H.W. planned and J.A.R., H.A., U.N., W.W.-L.W., L.G., B.G., S.M.C. and E.R. performed the analyses of cells, tissues and blood samples obtained from all mouse strains used in this study. H.W. and J.S. wrote the manuscript assisted by B.G., S.M.C., A.C.S., C.H.E., E.R. and A.I.W.

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Correspondence to Henning Walczak.

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Competing interests

H.W. is founder, shareholder and scientific advisor of Apogenix GmbH, Heidelberg, Germany. J.S. is a consultant for TetraLogic Pharmaceuticals, Malvern, Pennsylvania, USA.

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Gerlach, B., Cordier, S., Schmukle, A. et al. Linear ubiquitination prevents inflammation and regulates immune signalling. Nature 471, 591–596 (2011). https://doi.org/10.1038/nature09816

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