Nedd4 augments the adaptive immune response by promoting ubiquitin-mediated degradation of Cbl-b in activated T cells


Nedd4 and Itch are E3 ubiquitin ligases that ubiquitinate similar targets in vitro and thus are thought to function similarly. T cells lacking Itch show spontaneous activation and T helper type 2 polarization. To test whether loss of Nedd4 affects T cells in the same way, we generated Nedd4+/+ and Nedd4−/− fetal liver chimeras. Nedd4−/− T cells developed normally but proliferated less, produced less interleukin 2 and provided inadequate help to B cells. Nedd4−/− T cells contained more of the E3 ubiquitin ligase Cbl-b, and Nedd4 was required for polyubiquitination of Cbl-b induced by CD28 costimulation. Our data demonstrate that Nedd4 promotes the conversion of naive T cells into activated T cells. We propose that Nedd4 and Itch ubiquitinate distinct target proteins in vivo.

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Figure 1: Phenotypic analysis of Nedd4+/+ and Nedd4−/− cells isolated from thymus, lymph node and spleen.
Figure 2: Fewer Nedd4−/− T cells than Nedd4+/+ T cells express markers characteristic of an effector phenotype.
Figure 3: T cells lacking Nedd4 expand poorly in response to antigen in vivo and do not provide adequate help to B cells.
Figure 4: Fewer Nedd4−/− T cells than Nedd4+/+ T cells produce IL-2.
Figure 5: Nedd4−/− CD4+ T cells express normal amounts of PTEN and have higher expression of Cbl-b.
Figure 6: Nedd4 binds to and promotes the ubiquitination of Cbl-b.
Figure 7: Depletion of Cbl-b restores IL-2 production in Nedd4-depleted CD4+ T cells.


  1. 1

    Ciechanover, A., Heller, H., Elias, S., Haas, A.L. & Hershko, A. ATP-dependent conjugation of reticulocyte proteins with the polypeptide required for protein degradation. Proc. Natl. Acad. Sci. USA 77, 1365–1368 (1980).

    CAS  Article  Google Scholar 

  2. 2

    Ciechanover, A. Intracellular protein degradation: from a vague idea thru the lysosome and the ubiquitin-proteasome system and onto human diseases and drug targeting. Cell Death Differ. 12, 1178–1190 (2005).

    CAS  Article  Google Scholar 

  3. 3

    Liu, Y.C. Ubiquitin ligases and the immune response. Annu. Rev. Immunol. 22, 81–127 (2004).

    Article  Google Scholar 

  4. 4

    Fang, D. et al. Dysregulation of T lymphocyte function in itchy mice: a role for Itch in TH2 differentiation. Nat. Immunol. 3, 281–287 (2002).

    CAS  Article  Google Scholar 

  5. 5

    Oliver, P.M. et al. Ndfip1 protein promotes the function of itch ubiquitin ligase to prevent T cell activation and T helper 2 cell-mediated inflammation. Immunity 25, 929–940 (2006).

    CAS  Article  Google Scholar 

  6. 6

    Heissmeyer, V. et al. Calcineurin imposes T cell unresponsiveness through targeted proteolysis of signaling proteins. Nat. Immunol. 5, 255–265 (2004).

    CAS  Article  Google Scholar 

  7. 7

    Babu, S., Blauvelt, C.P., Kumaraswami, V. & Nutman, T.B. Regulatory networks induced by live parasites impair both Th1 and Th2 pathways in patent lymphatic filariasis: implications for parasite persistence. J. Immunol. 176, 3248–3256 (2006).

    CAS  Article  Google Scholar 

  8. 8

    Perry, W.L. et al. The itchy locus encodes a novel ubiquitin protein ligase that is disrupted in a18H mice. Nat. Genet. 18, 143–146 (1998).

    CAS  Article  Google Scholar 

  9. 9

    Liu, Y.C. The E3 ubiquitin ligase Itch in T cell activation, differentiation, and tolerance. Semin. Immunol. 19, 197–205 (2007).

    CAS  Article  Google Scholar 

  10. 10

    Magnifico, A. et al. WW domain HECT E3s target Cbl RING finger E3s for proteasomal degradation. J. Biol. Chem. 278, 43169–43177 (2003).

    CAS  Article  Google Scholar 

  11. 11

    Scharschmidt, E., Wegener, E., Heissmeyer, V., Rao, A. & Krappmann, D. Degradation of Bcl10 induced by T-cell activation negatively regulates NF-κB signaling. Mol. Cell. Biol. 24, 3860–3873 (2004).

    CAS  Article  Google Scholar 

  12. 12

    Bonnevier, J.L., Zhang, R. & Mueller, D.L. E3 ubiquitin ligases and their control of T cell autoreactivity. Arthritis Res. Ther. 7, 233–242 (2005).

    CAS  Article  Google Scholar 

  13. 13

    Mueller, D.L. E3 ubiquitin ligases as T cell anergy factors. Nat. Immunol. 5, 883–890 (2004).

    CAS  Article  Google Scholar 

  14. 14

    Ingham, R.J. et al. WW domains provide a platform for the assembly of multiprotein networks. Mol. Cell. Biol. 25, 7092–7106 (2005).

    CAS  Article  Google Scholar 

  15. 15

    Stryke, D. et al. BayGenomics: a resource of insertional mutations in mouse embryonic stem cells. Nucleic Acids Res. 31, 278–281 (2003).

    CAS  Article  Google Scholar 

  16. 16

    Rees, W. et al. An inverse relationship between T cell receptor affinity and antigen dose during CD4+ T cell responses in vivo and in vitro. Proc. Natl. Acad. Sci. USA 96, 9781–9786 (1999).

    CAS  Article  Google Scholar 

  17. 17

    Grewal, I.S. & Flavell, R.A. The role of CD40 ligand in costimulation and T-cell activation. Immunol. Rev. 153, 85–106 (1996).

    CAS  Article  Google Scholar 

  18. 18

    Powell, J.D., Ragheb, J.A., Kitagawa-Sakakida, S. & Schwartz, R.H. Molecular regulation of interleukin-2 expression by CD28 co-stimulation and anergy. Immunol. Rev. 165, 287–300 (1998).

    CAS  Article  Google Scholar 

  19. 19

    Umlauf, S.W., Beverly, B., Lantz, O. & Schwartz, R.H. Regulation of interleukin 2 gene expression by CD28 costimulation in mouse T-cell clones: both nuclear and cytoplasmic RNAs are regulated with complex kinetics. Mol. Cell. Biol. 15, 3197–3205 (1995).

    CAS  Article  Google Scholar 

  20. 20

    Green, J.M. et al. Absence of B7-dependent responses in CD28-deficient mice. Immunity 1, 501–508 (1994).

    CAS  Article  Google Scholar 

  21. 21

    Lucas, P.J., Negishi, I., Nakayama, K., Fields, L.E. & Loh, D.Y. Naive CD28-deficient T cells can initiate but not sustain an in vitro antigen-specific immune response. J. Immunol. 154, 5757–5768 (1995).

    CAS  PubMed  Google Scholar 

  22. 22

    Shahinian, A. et al. Differential T cell costimulatory requirements in CD28-deficient mice. Science 261, 609–612 (1993).

    CAS  Article  Google Scholar 

  23. 23

    Trotman, L.C. et al. Ubiquitination regulates PTEN nuclear import and tumor suppression. Cell 128, 141–156 (2007).

    CAS  Article  Google Scholar 

  24. 24

    Wang, X. et al. NEDD4 is a proto-oncogenic ubiquitin ligase for PTEN. Cell 128, 129–139 (2007).

    CAS  Article  Google Scholar 

  25. 25

    Leykauf, K. et al. Ubiquitin protein ligase Nedd4 binds to connexin43 by a phosphorylation-modulated process. J. Cell Sci. 119, 3634–3642 (2006).

    CAS  Article  Google Scholar 

  26. 26

    Seminario, M.C. & Wange, R.L. Lipid phosphatases in the regulation of T cell activation: living up to their PTEN-tial. Immunol. Rev. 192, 80–97 (2003).

    CAS  Article  Google Scholar 

  27. 27

    Xu, Z., Stokoe, D., Kane, L.P. & Weiss, A. The inducible expression of the tumor suppressor gene PTEN promotes apoptosis and decreases cell size by inhibiting the PI3K/Akt pathway in Jurkat T cells. Cell Growth Differ. 13, 285–296 (2002).

    CAS  PubMed  Google Scholar 

  28. 28

    Buckler, J.L., Walsh, P.T., Porrett, P.M., Choi, Y. & Turka, L.A. Cutting edge: T cell requirement for CD28 costimulation is due to negative regulation of TCR signals by PTEN. J. Immunol. 177, 4262–4266 (2006).

    CAS  Article  Google Scholar 

  29. 29

    Zhang, J. et al. Cutting edge: regulation of T cell activation threshold by CD28 costimulation through targeting Cbl-b for ubiquitination. J. Immunol. 169, 2236–2240 (2002).

    CAS  Article  Google Scholar 

  30. 30

    Kim, H.T. et al. Certain pairs of ubiquitin-conjugating enzymes (E2s) and ubiquitin-protein ligases (E3s) synthesize nondegradable forked ubiquitin chains containing all possible isopeptide linkages. J. Biol. Chem. 282, 17375–17386 (2007).

    CAS  Article  Google Scholar 

  31. 31

    Levy, F. et al. Ubiquitylation of a melanosomal protein by HECT-E3 ligases serves as sorting signal for lysosomal degradation. Mol. Biol. Cell 16, 1777–1787 (2005).

    CAS  Article  Google Scholar 

  32. 32

    Murdaca, J. et al. Grb10 prevents Nedd4-mediated vascular endothelial growth factor receptor-2 degradation. J. Biol. Chem. 279, 26754–26761 (2004).

    CAS  Article  Google Scholar 

  33. 33

    Vecchione, A., Marchese, A., Henry, P., Rotin, D. & Morrione, A. The Grb10/Nedd4 complex regulates ligand-induced ubiquitination and stability of the insulin-like growth factor I receptor. Mol. Cell. Biol. 23, 3363–3372 (2003).

    CAS  Article  Google Scholar 

  34. 34

    Sakata, T. et al. Drosophila Nedd4 regulates endocytosis of Notch and suppresses its ligand-independent activation. Curr. Biol. 14, 2228–2236 (2004).

    CAS  Article  Google Scholar 

  35. 35

    Jennings, M.D., Blankley, R.T., Baron, M., Golovanov, A.P. & Avis, J.M. Specificity and autoregulation of Notch binding by tandem WW domains in suppressor of Deltex. J. Biol. Chem. 282, 29032–29042 (2007).

    CAS  Article  Google Scholar 

  36. 36

    Chiang, Y.J. et al. Cbl-b regulates the CD28 dependence of T-cell activation. Nature 403, 216–220 (2000).

    CAS  Article  Google Scholar 

  37. 37

    Di Cristofano, A. et al. Impaired Fas response and autoimmunity in Pten+/− mice. Science 285, 2122–2125 (1999).

    CAS  Article  Google Scholar 

  38. 38

    Donovan, J.A., Wange, R.L., Langdon, W.Y. & Samelson, L.E. The protein product of the c–cbl protooncogene is the 120-kDa tyrosine-phosphorylated protein in Jurkat cells activated via the T cell antigen receptor. J. Biol. Chem. 269, 22921–22924 (1994).

    CAS  PubMed  Google Scholar 

  39. 39

    Gaide, O. et al. CARMA1 is a critical lipid raft-associated regulator of TCR-induced NF-κB activation. Nat. Immunol. 3, 836–843 (2002).

    CAS  Article  Google Scholar 

  40. 40

    Robey, E. et al. An activated form of Notch influences the choice between CD4 and CD8 T cell lineages. Cell 87, 483–492 (1996).

    CAS  Article  Google Scholar 

  41. 41

    Thome, M. & Tschopp, J. TCR-induced NF-κB activation: a crucial role for Carma1, Bcl10 and MALT1. Trends Immunol. 24, 419–424 (2003).

    CAS  Article  Google Scholar 

  42. 42

    Villalba, M. et al. Protein kinase cθ cooperates with calcineurin to induce Fas ligand expression during activation-induced T cell death. J. Immunol. 163, 5813–5819 (1999).

    CAS  PubMed  Google Scholar 

  43. 43

    Washburn, T. et al. Notch activity influences the αβ versus γδ T cell lineage decision. Cell 88, 833–843 (1997).

    CAS  Article  Google Scholar 

  44. 44

    McDonald, F.J. et al. Disruption of the β subunit of the epithelial Na+ channel in mice: hyperkalemia and neonatal death associated with a pseudohypoaldosteronism phenotype. Proc. Natl. Acad. Sci. USA 96, 1727–1731 (1999).

    CAS  Article  Google Scholar 

  45. 45

    Cao, X.R. et al. Nedd4 controls animal growth by regulating IGF-1 signaling. Sci. Signal. (in the press).

  46. 46

    McCormack, J.E., Kappler, J. & Marrack, P. Stimulation with specific antigen can block superantigen-mediated deletion of T cells in vivo. Proc. Natl. Acad. Sci. USA 91, 2086–2090 (1994).

    CAS  Article  Google Scholar 

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We thank R. Schwartz (National Institute of Allergy and Infectious Diseases) for Cbl-b-deficient mice; M. Shaffer for help with siRNA experiments; and J. Loomis for sorting cells.

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Correspondence to Paula M Oliver.

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Yang, B., Gay, D., MacLeod, M. et al. Nedd4 augments the adaptive immune response by promoting ubiquitin-mediated degradation of Cbl-b in activated T cells. Nat Immunol 9, 1356–1363 (2008).

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