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:

ABIN-1 regulates RIPK1 activation by linking Met1 ubiquitylation with Lys63 deubiquitylation in TNF-RSC

Nature Cell Biology volume 20, pages 58–68 (2018)Cite this article

Abstract

Ubiquitylation of the TNFR1 signalling complex (TNF-RSC) controls the activation of RIPK1, a kinase critically involved in mediating multiple TNFα-activated deleterious events. However, the molecular mechanism that coordinates different types of ubiquitylation modification to regulate the activation of RIPK1 kinase remains unclear. Here, we show that ABIN-1/NAF-1, a ubiquitin-binding protein, is recruited rapidly into TNF-RSC in a manner dependent on the Met1-ubiquitylating complex LUBAC to regulate the recruitment of A20 to control Lys63 deubiquitylation of RIPK1. ABIN-1 deficiency reduces the recruitment of A20 and licenses cells to die through necroptosis by promoting Lys63 ubiquitylation and activation of RIPK1 with TNFα stimulation under conditions that would otherwise exclusively activate apoptosis in wild-type cells. Inhibition of RIPK1 kinase and RIPK3 deficiency block the embryonic lethality of Abin-1 –/– mice. We propose that ABIN-1 provides a critical link between Met1 ubiquitylation mediated by the LUBAC complex and Lys63 deubiquitylation by phospho-A20 to modulate the activation of RIPK1.

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

Fig. 1: ABIN-1 deficiency sensitizes cells to necroptosis.
Fig. 2: Elevated biochemical hallmarks of necroptosis in Abin-1 –/– cells stimulated by TNFα/CHX/zVAD.fmk.
Fig. 3: Inhibition of RIPK1, RIPK3 and MLKL prolongs the survival of apoptotic ABIN-1-deficient cells induced by TNFα/CHX.
Fig. 4: Inhibition of necroptosis prolongs cell survival but has no effect on caspase activation in Abin-1 –/– cells treated with TNFα/CHX.
Fig. 5: Increased Lys63 ubiquitylation of RIPK1 in Abin-1 –/– MEFs stimulated by TNFα.
Fig. 6: ABIN-1 is important for the recruitment of p-A20 into TNF-RSC.
Fig. 7: The presence of hallmarks of both apoptosis and necroptosis in Abin-1 –/– embryos.

Similar content being viewed by others

References

  1. Ofengeim, D. & Yuan, J. Regulation of RIP1 kinase signalling at the crossroads of inflammation and cell death. Nat. Rev. Mol. Cell Biol. 14, 727–736 (2013).

    Article  CAS  PubMed  Google Scholar 

  2. Wallach, D., Kang, T. B., Dillon, C. P. & Green, D. R. Programmed necrosis in inflammation: toward identification of the effector molecules. Science 352, aaf2154 (2016).

    Article  PubMed  Google Scholar 

  3. Cho, Y. S. et al. Phosphorylation-driven assembly of the RIP1-RIP3 complex regulates programmed necrosis and virus-induced inflammation. Cell 137, 1112–1123 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. He, S. et al. Receptor interacting protein kinase-3 determines cellular necrotic response toTNF-a. Cell 137, 1100–1111 (2009).

    Article  CAS  PubMed  Google Scholar 

  5. Degterev, A. et al. Identification of RIP1 kinase as a specific cellular target of necrostatins. Nat. Chem. Biol. 4, 313–321 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Vucic, D., Dixit, V. M. & Wertz, I. E. Ubiquitylation in apoptosis: a post-translational modification at the edge of life and death. Nat. Rev. Mol. Cell Biol. 12, 439–452 (2011).

    Article  CAS  PubMed  Google Scholar 

  7. Ito, Y. et al. RIPK1 mediates axonal degeneration by promoting inflammation and necroptosis in ALS. Science 353, 603–608 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Pasparakis, M. & Vandenabeele, P. Necroptosis and its role in inflammation. Nature 517, 311–320 (2015).

    Article  CAS  PubMed  Google Scholar 

  9. Welz, P. S. et al. FADD prevents RIP3-mediated epithelial cell necrosis and chronic intestinal inflammation. Nature 477, 330–334 (2011).

    Article  CAS  PubMed  Google Scholar 

  10. Vlantis, K. et al. NEMO prevents RIP kinase 1-mediated epithelial cell death and chronic intestinal inflammation by NF-κB-dependent and -independent functions. Immunity 44, 553–567 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Harris, P. A. et al. DNA-encoded library screening identifies benzo[b][1,4]oxazepin-4-ones as highly potent and monoselective receptor interacting protein 1 kinase inhibitors. J. Med. Chem. 59, 2163–2178 (2016).

    Article  CAS  PubMed  Google Scholar 

  12. Matmati, M. et al. A20 (TNFAIP3) deficiency in myeloid cells triggers erosive polyarthritis resembling rheumatoid arthritis. Nat. Genet. 43, 908–912 (2011).

    Article  CAS  PubMed  Google Scholar 

  13. Catrysse, L., Vereecke, L., Beyaert, R. & van Loo, G. A20 in inflammation and autoimmunity. Trends Immunol. 35, 22–31 (2014).

    Article  CAS  PubMed  Google Scholar 

  14. Hsu, H., Huang, J., Shu, H. B., Baichwal, V. & Goeddel, D. V. TNF-dependent recruitment of the protein kinase RIP to the TNF receptor-1 signaling complex. Immunity 4, 387–396 (1996).

    Article  CAS  PubMed  Google Scholar 

  15. Dondelinger, Y., Darding, M., Bertrand, M. J. & Walczak, H. Poly-ubiquitination in TNFR1-mediated necroptosis. Cell. Mol. Life Sci. 73, 2165–2176 (2016).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Bertrand, M. J. et al. cIAP1 and cIAP2 facilitate cancer cell survival by functioning as E3 ligases that promote RIP1 ubiquitination. Mol. Cell 30, 689–700 (2008).

    Article  CAS  PubMed  Google Scholar 

  17. 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  PubMed  Google Scholar 

  18. Gerlach, B. et al. Linear ubiquitination prevents inflammation and regulates immune signalling. Nature 471, 591–596 (2011).

    Article  CAS  PubMed  Google Scholar 

  19. Degterev, A. et al. Chemical inhibitor of nonapoptotic cell death with therapeutic potential for ischemic brain injury. Nat. Chem. Biol. 1, 112–119 (2005).

    Article  CAS  PubMed  Google Scholar 

  20. Nanda, S. K. et al. Polyubiquitin binding to ABIN1 is required to prevent autoimmunity. J. Exp. Med. 208, 1215–1228 (2011).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  21. Callahan, J. A. et al. Cutting edge: ABIN-1 protects against psoriasis by restricting MyD88 signals in dendritic cells. J. Immunol. 191, 535–539 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Caster, D. J. et al. ABIN1 dysfunction as a genetic basis for lupus nephritis. J. Am. Soc. Nephrol. 24, 1743–1754 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  23. Oshima, S. et al. ABIN-1 is a ubiquitin sensor that restricts cell death and sustains embryonic development. Nature 457, 906–909 (2009).

    Article  CAS  PubMed  Google Scholar 

  24. Lu, J. et al. SM-164: a novel, bivalent Smac mimetic that induces apoptosis and tumor regression by concurrent removal of the blockade of cIAP-1/2 and XIAP. Cancer Res. 68, 9384–9393 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  25. Polykratis, A. et al. Cutting edge: RIPK1 kinase inactive mice are viable and protected from TNF-induced necroptosis in vivo. J. Immunol. 193, 1539–1543 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Newton, K., Sun, X. & Dixit, V. M. Kinase RIP3 is dispensable for normal NF-κBs, signaling by the B-cell and T-cell receptors, tumor necrosis factor receptor 1, and Toll-like receptors 2 and 4. Mol. Cell. Biol. 24, 1464–1469 (2004).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Ofengeim, D. et al. Activation of necroptosis in multiple sclerosis. Cell Rep. 10, 1836–1849 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Berger, S. B. et al. Cutting edge: RIP1 kinase activity is dispensable for normal development but is a key regulator of inflammation in SHARPIN-deficient mice. J. Immunol. 192, 5476–5480 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  29. Zhang, D. W. et al. RIP3, an energy metabolism regulator that switches TNF-induced cell death from apoptosis to necrosis. Science 325, 332–336 (2009).

    Article  CAS  PubMed  Google Scholar 

  30. Sun, L. et al. Mixed lineage kinase domain-like protein mediates necrosis signaling downstream of RIP3 kinase. Cell 148, 213–227 (2012).

    Article  CAS  PubMed  Google Scholar 

  31. Dondelinger, Y. et al. NF-κB-independent role of IKKα/IKKβ in preventing RIPK1 kinase-dependent apoptotic and necroptotic cell death during TNF signaling. Mol. Cell 60, 63–76 (2015).

    Article  CAS  PubMed  Google Scholar 

  32. Mihaly, S. R., Ninomiya-Tsuji, J. & Morioka, S. TAK1 control of cell death. Cell Death Differ. 21, 1667–1676 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Wang, L., Du, F. & Wang, X. TNF-α induces two distinct caspase-8 activation pathways. Cell 133, 693–703 (2008).

    Article  CAS  PubMed  Google Scholar 

  34. Lin, Y., Devin, A., Rodriguez, Y. & Liu, Z. G. Cleavage of the death domain kinase RIP by caspase-8 prompts TNF-induced apoptosis. Genes Dev. 13, 2514–2526 (1999).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  35. Li, J. et al. The RIP1/RIP3 necrosome forms a functional amyloid signaling complex required for programmed necrosis. Cell 150, 339–350 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Wertz, I. E. et al. Phosphorylation and linear ubiquitin direct A20 inhibition of inflammation. Nature 528, 370–375 (2015).

    Article  CAS  PubMed  Google Scholar 

  37. Newton, K. et al. Ubiquitin chain editing revealed by polyubiquitin linkage-specific antibodies. Cell 134, 668–678 (2008).

    Article  CAS  PubMed  Google Scholar 

  38. Draber, P. et al. LUBAC-recruited CYLD and A20 regulate gene activation and cell death by exerting opposing effects on linear ubiquitin in signaling complexes. Cell Rep. 13, 2258–2272 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Reiley, W., Zhang, M., Wu, X., Granger, E. & Sun, S. C. Regulation of the deubiqui-tinating enzyme CYLD by IκB kinase gamma-dependent phosphorylation. Mol. Cell. Biol. 25, 3886–3895 (2005).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Hutti, J. E. et al. IkB kinase p phosphorylates the K63 deubiquitinase A20 to cause feedback inhibition of the NF-κB pathway. Mol. Cell. Biol. 27, 7451–7461 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  41. Raingeaud, J. et al. Pro-inflammatory cytokines and environmental stress cause p38 mitogen-activated protein kinase activation by dual phosphorylation on tyrosine and threonine. J. Biol. Chem. 270, 7420–7426 (1995).

    Article  CAS  PubMed  Google Scholar 

  42. Hutti, J. E. et al. Phosphorylation of the tumor suppressor CYLD by the breast cancer oncogene IKKε promotes cell transformation. Mol. Cell 34, 461–472 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Heyninck, K., Kreike, M. M. & Beyaert, R. Structure-function analysis of the A20-binding inhibitor of NF-κB activation, ABIN-1. FEBS Lett. 536, 135–140 (2003).

    Article  CAS  PubMed  Google Scholar 

  44. Dynek, J. N. et al. c-IAP1 andUbcH5promoteK11-linked polyubiquitination of RIP1 in TNF signalling. EMBO J. 29, 4198–4209 (2010).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. Bosanac, I. et al. Ubiquitin binding to A20 ZnF4 is required for modulation of NF-κB signaling. Mol. Cell 40, 548–557 (2010).

    Article  CAS  PubMed  Google Scholar 

  46. Lee, E. G. et al. Failure to regulate TNF-induced NF-κB and cell death responses in A20-deficient mice. Science 289, 2350–2354 (2000).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Onizawa, M. et al. The ubiquitin-modifying enzyme A20 restricts ubiquitination of the kinase RIPK3 and protects cells from necroptosis. Nat. Immunol. 16, 618–627 (2015).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  48. Yi, C. H. & Yuan, J. The Jekyll and Hyde functions of caspases. Dev. Cell 16, 21–34 (2009).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  49. Dillon, C. P. et al. Survival function of the FADD-CASPASE-8-cFLIP(L) complex. Cell Rep. 1, 401–407 (2012).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  50. Zhang, J., Cado, D., Chen, A., Kabra, N. H. & Winoto, A. Fas-mediated apoptosis and activation-induced T-cell proliferation are defective in mice lackingFADD/Mort1. Nature 392, 296–300 (1998).

    Article  CAS  PubMed  Google Scholar 

  51. Yeh, W. C. et al. Requirement for Casper (c-FLIP) in regulation of death receptor-induced apoptosis and embryonic development. Immunity 12, 633–642 (2000).

    Article  CAS  PubMed  Google Scholar 

  52. Varfolomeev, E. E. et al. Targeted disruption of the mouse Caspase 8 gene ablates cell death induction by the TNF receptors, Fas/Apo1, and DR3 and is lethal prenatally. Immunity 9, 267–276 (1998).

    Article  CAS  PubMed  Google Scholar 

  53. Xu, L. et al. c-IAP1 cooperates with Myc by acting as a ubiquitin ligase for Mad1. Mol. Cell 28, 914–922 (2007).

    Article  CAS  PubMed  Google Scholar 

  54. Lu, J. et al. SM-164: a novel, bivalent Smac mimetic that induces apoptosis and tumor regression by concurrent removal of the blockade of cIAP-1/2 and XIAP. Cancer Res. 68, 9384–9393 (2008).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

This work was supported in part by grants (to J.Y.) from the NINDS (1R01NS082257) and the NIA (1R01AG047231) and from the Chinese Academy of Sciences, the China Ministry of Science and Technology Program (2014ZX09102001-002), the China National Natural Science Foundation (31530041), the National Key R&D Program of China, and the National Key Research and Development Program (2016YFA0501900). We thank V. Dixit of Genentech for the Ripk3 –/– mice and for the antibody against Lys63 ubiquitin chains, M. Pasparakis of University of Cologne, Germany, and M. A. Kelliher of University of Massachusetts, USA, for providing the Ripk1 D138N mice, and H. Walczak of Imperial College, UK, for providing the Hoip –/– MEFs. We thank G. Kasof of Cell Signaling for generating the p-Ser166 antibodyand R. Bronson for mouse histopathology analysis. We thank D. Ofengeim and A. D. Yu for comments on the manuscript and the members of the Yuan laboratory for stimulating discussions.

Author information

Authors and Affiliations

  1. Department of Cell Biology, Harvard Medical School, Boston, MA, USA

    Slawomir A. Dziedzic, Zhenyi Su, Vica Jean Barrett, Ayaz Najafov, Adnan K. Mookhtiar, Palak Amin, Hong Zhu & Junying Yuan

  2. Department of Biochemistry and Molecular Biology, Medical School, Southeast University, Nanjing, Jiangsu, China

    Zhenyi Su

  3. Interdisciplinary Research Center on Biology and Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Science, PuDong District, Shanghai, China

    Heling Pan, Li Sun & Junying Yuan

  4. Department of Medicine, University of California, San Francisco, San Francisco, CA, USA

    Averil Ma

  5. Department of Pathology, Case Western Reserve University School of Medicine, Cleveland, OH, USA

    Derek W. Abbott

Authors
  1. Slawomir A. Dziedzic
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Zhenyi Su
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Vica Jean Barrett
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Ayaz Najafov
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Adnan K. Mookhtiar
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Palak Amin
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Heling Pan
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Li Sun
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Hong Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Averil Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. Derek W. Abbott
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. Junying Yuan
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.Y. conceived the concept, designed the experiments and wrote the manuscript. S.A.D. designed the experiments, executed the majority of the experiments and prepared the figures. Z.S., V.J.B., A.N., A.K.M., P.A., H.P. and L.S. conducted specific experiments. D.W.A. provided the p-S381 A20 antibody, A20 plasmids and A20-mutant-complemented cells. A.M. provided Abin-1 –/+ mice. H.Z. and V.J.B. performed genotyping.

Corresponding author

Correspondence to Junying Yuan.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Additional information

Publisher’s note: Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Electronic supplementary material

Supplementary Information

Supplementary Figures 1-6 and Supplementary Figure and Table legends.

Life Sciences Reporting Summary

Supplementary Table 1

The embryonic lethality of Abin-1 -/- mice is rescued by inactivation of RIPK1 kinase activity in Abin-1 -/-; Ripk1 D138N/D138N mice and RIPK3 deficiency in Abin-1 -/-; Ripk3 -/- mice. Abin-1 +/-; Ripk1 D138N/+ males and females were mated to obtain 144 pups; and Abin-1 +/-; Ripk3 +/- males and females were mated to obtain 160 pups as described in the table

Supplementary Table 2

Statistics source data. Source data of all repeats for Fig. 1, Fig. 3, Supplementary Fig. 1a,b, Supplementary Fig. 2a, Supplementary Fig. 4c and Supplementary Fig. 5 accompanied by precise P values (where applicable) are provided as different sheets.

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Dziedzic, S.A., Su, Z., Jean Barrett, V. et al. ABIN-1 regulates RIPK1 activation by linking Met1 ubiquitylation with Lys63 deubiquitylation in TNF-RSC. Nat Cell Biol 20, 58–68 (2018). https://doi.org/10.1038/s41556-017-0003-1

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/s41556-017-0003-1

This article is cited by

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