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:

Scaffold protein Dlgh1 coordinates alternative p38 kinase activation, directing T cell receptor signals toward NFAT but not NF-κB transcription factors

Nature Immunology volume 8, pages 154–161 (2007)Cite this article

Abstract

Tyrosine kinases couple the T cell receptor (TCR) to discrete signaling cascades, each of which is capable of inducing a distinct functional outcome. Precisely how TCR signals are channeled toward specific targets remains unclear. TCR stimulation triggers 'alternative' activation of the mitogen-activated protein kinase p38, whereby the Lck and Zap70 tyrosine kinases directly activate p38. Here we report that alternatively activated p38 associated with the Dlgh1 MAGUK scaffold protein. 'Knockdown' of Dlgh1 expression blocked TCR-induced activation of p38 and the transcription factor NFAT but not of the mitogen-activated protein kinase Jnk or transcription factor NF-κB. A Dlgh1 mutant incapable of binding p38 failed to activate NFAT. Along with reports that the CARMA1 MAGUK scaffold protein coordinates activation of Jnk and NF-κB but not of p38 or NFAT, our findings identify MAGUK scaffold proteins as 'orchestrators' of TCR signal specificity.

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: The kinase p38 binds to Dlgh1 PDZ domains and is phosphorylated after TCR and CD28 stimulation.
Figure 2: Diminished Dlgh1 expression impairs alternative activation of p38 but not of Jnk or Erk.
Figure 3: Dlgh1 facilitates activation of NFAT but not of NF-κB.
Figure 4: Activity by p38 is required for Dlgh1-mediated NFAT activation.
Figure 5: Disruption of the Dlgh1-p38 interaction impairs NFAT and alternative p38 activation and IFN-γ production.

Similar content being viewed by others

References

  1. Elion, E.A., Qi, M. & Chen, W. Signal transduction. Signaling specificity in yeast. Science 307, 687–688 (2005).

    Article  CAS  Google Scholar 

  2. Hoshi, N., Langeberg, L.K. & Scott, J.D. Distinct enzyme combinations in AKAP signalling complexes permit functional diversity. Nat. Cell Biol. 7, 1066–1073 (2005).

    Article  CAS  Google Scholar 

  3. Ptashne, M. & Gann, A. Signal transduction. Imposing specificity on kinases. Science 299, 1025–1027 (2003).

    Article  CAS  Google Scholar 

  4. Burack, W.R. & Shaw, A.S. Signal transduction: hanging on a scaffold. Curr. Opin. Cell Biol. 12, 211–216 (2000).

    Article  CAS  Google Scholar 

  5. Jordan, M.S., Singer, A.L. & Koretzky, G.A. Adaptors as central mediators of signal transduction in immune cells. Nat. Immunol. 4, 110–116 (2003).

    Article  CAS  Google Scholar 

  6. Round, J.L. et al. Dlgh1 coordinates actin polymerization, synaptic T cell receptor and lipid raft aggregation, and effector function in T cells. J. Exp. Med. 201, 419–430 (2005).

    Article  CAS  Google Scholar 

  7. Ludford-Menting, M.J. et al. A network of PDZ-containing proteins regulates T cell polarity and morphology during migration and immunological synapse formation. Immunity 22, 737–748 (2005).

    Article  CAS  Google Scholar 

  8. da Silva, A.J. et al. Cloning of a novel T-cell protein FYB that binds FYN and SH2-domain-containing leukocyte protein 76 and modulates interleukin 2 production. Proc. Natl. Acad. Sci. USA 94, 7493–7498 (1997).

    Article  CAS  Google Scholar 

  9. Abbas, A.K. & Sen, R. The activation of lymphocytes is in their CARMA. Immunity 18, 721–722 (2003).

    Article  CAS  Google Scholar 

  10. Geng, L. & Rudd, C.E. Signalling scaffolds and adaptors in T-cell immunity. Br. J. Haematol. 116, 19–27 (2002).

    Article  CAS  Google Scholar 

  11. Im, S.H. & Rao, A. Activation and deactivation of gene expression by Ca2+/calcineurin-NFAT-mediated signaling. Mol. Cells 18, 1–9 (2004).

    CAS  PubMed  Google Scholar 

  12. Serfling, E. et al. NFAT and NF-κB factors-the distant relatives. Int. J. Biochem. Cell Biol. 36, 1166–1170 (2004).

    Article  CAS  Google Scholar 

  13. Herndon, T.M., Shan, X.C., Tsokos, G.C. & Wange, R.L. ZAP-70 and SLP-76 regulate protein kinase C-θ and NF-κB activation in response to engagement of CD3 and CD28. J. Immunol. 166, 5654–5664 (2001).

    Article  CAS  Google Scholar 

  14. Macian, F. NFAT proteins: key regulators of T-cell development and function. Nat. Rev. Immunol. 5, 472–484 (2005).

    Article  CAS  Google Scholar 

  15. Cianferoni, A. et al. Defective nuclear translocation of nuclear factor of activated T cells and extracellular signal-regulated kinase underlies deficient IL-2 gene expression in Wiskott-Aldrich syndrome. J. Allergy Clin. Immunol. 116, 1364–1371 (2005).

    Article  CAS  Google Scholar 

  16. Rudd, C.E. MAPK p38: alternative and nonstressful in T cells. Nat. Immunol. 6, 368–370 (2005).

    Article  CAS  Google Scholar 

  17. Rincon, M. & Pedraza-Alva, G. JNK and p38 MAP kinases in CD4+ and CD8+ T cells. Immunol. Rev. 192, 131–142 (2003).

    Article  CAS  Google Scholar 

  18. Rincon, M., Flavell, R.A. & Davis, R.A. The JNK and P38 MAP kinase signaling pathways in T cell-mediated immune responses. Free Radic. Biol. Med. 28, 1328–1337 (2000).

    Article  CAS  Google Scholar 

  19. Salvador, J.M. et al. Alternative p38 activation pathway mediated by T cell receptor–proximal tyrosine kinases. Nat. Immunol. 6, 390–395 (2005).

    Article  CAS  Google Scholar 

  20. Salvador, J.M. et al. The autoimmune suppressor Gadd45α inhibits the T cell alternative p38 activation pathway. Nat. Immunol. 6, 396–402 (2005).

    Article  CAS  Google Scholar 

  21. Hanada, T. et al. Human homologue of the Drosophila discs large tumor suppressor binds to p56lck tyrosine kinase and Shaker type Kv1.3 potassium channel in T lymphocytes. J. Biol. Chem. 272, 26899–26904 (1997).

    Article  CAS  Google Scholar 

  22. Xavier, R. et al. Discs large (Dlg1) complexes in lymphocyte activation. J. Cell Biol. 166, 173–178 (2004).

    Article  CAS  Google Scholar 

  23. Jun, J.E. et al. Identifying the MAGUK protein Carma-1 as a central regulator of humoral immune responses and atopy by genome-wide mouse mutagenesis. Immunity 18, 751–762 (2003).

    Article  CAS  Google Scholar 

  24. Hara, H. et al. The MAGUK family protein CARD11 is essential for lymphocyte activation. Immunity 18, 763–775 (2003).

    Article  CAS  Google Scholar 

  25. Wang, D. et al. CD3/CD28 costimulation-induced NF-κB activation is mediated by recruitment of protein kinase C-θ, Bcl10, and IκB kinase β to the immunological synapse through CARMA1. Mol. Cell. Biol. 24, 164–171 (2004).

    Article  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  27. Pomerantz, J.L., Denny, E.M. & Baltimore, D. CARD11 mediates factor-specific activation of NF-κB by the T cell receptor complex. EMBO J. 21, 5184–5194 (2002).

    Article  CAS  Google Scholar 

  28. Miyoshi, J. & Takai, Y. Molecular perspective on tight-junction assembly and epithelial polarity. Adv. Drug Deliv. Rev. 57, 815–855 (2005).

    Article  CAS  Google Scholar 

  29. Dustin, M.L. & Colman, D.R. Neural and immunological synaptic relations. Science 298, 785–789 (2002).

    Article  CAS  Google Scholar 

  30. Dimitratos, S.D., Woods, D.F., Stathakis, D.G. & Bryant, P.J. Signaling pathways are focused at specialized regions of the plasma membrane by scaffolding proteins of the MAGUK family. Bioessays 21, 912–921 (1999).

    Article  CAS  Google Scholar 

  31. Kim, E. et al. Clustering of Shaker-type K+ channels by interaction with a family of membrane-associated guanylate kinases. Nature 378, 85–88 (1995).

    Article  CAS  Google Scholar 

  32. Okamura, H. et al. Concerted dephosphorylation of the transcription factor NFAT1 induces a conformational switch that regulates transcriptional activity. Mol. Cell 6, 539–550 (2000).

    Article  CAS  Google Scholar 

  33. Crabtree, G.R. & Olson, E.N. NFAT signaling: choreographing the social lives of cells. Cell 109, S67–S79 (2002).

    Article  CAS  Google Scholar 

  34. Zhou, B. et al. Regulation of the murine Nfatc1 gene by NFATc2. J. Biol. Chem. 277, 10704–10711 (2002).

    Article  CAS  Google Scholar 

  35. Attar, R.M. et al. Expression of constitutively active IκB beta in T cells of transgenic mice: persistent NF-κB activity is required for T-cell immune responses. Mol. Cell. Biol. 18, 477–487 (1998).

    Article  CAS  Google Scholar 

  36. Chiao, P.J., Miyamoto, S. & Verma, I.M. Autoregulation of IκBα activity. Proc. Natl. Acad. Sci. USA 91, 28–32 (1994).

    Article  CAS  Google Scholar 

  37. Yang, T.T. et al. Phosphorylation of NFATc4 by p38 mitogen-activated protein kinases. Mol. Cell. Biol. 22, 3892–3904 (2002).

    Article  CAS  Google Scholar 

  38. Gomez del Arco, P. et al. A role for the p38 MAP kinase pathway in the nuclear shuttling of NFATp. J. Biol. Chem. 275, 13872–13878 (2000).

    Article  CAS  Google Scholar 

  39. Wu, C.C., Hsu, S.C., Shih, H.M. & Lai, M.Z. Nuclear factor of activated T cells c is a target of p38 mitogen-activated protein kinase in T cells. Mol. Cell. Biol. 23, 6442–6454 (2003).

    Article  CAS  Google Scholar 

  40. Tavalin, S.J. et al. Regulation of GluR1 by the A-kinase anchoring protein 79 (AKAP79) signaling complex shares properties with long-term depression. J. Neurosci. 22, 3044–3051 (2002).

    Article  CAS  Google Scholar 

  41. Oliveria, S.F., Gomez, L.L. & Dell'Acqua, M.L. Imaging kinase–AKAP79–phosphatase scaffold complexes at the plasma membrane in living cells using FRET microscopy. J. Cell Biol. 160, 101–112 (2003).

    Article  CAS  Google Scholar 

  42. San-Antonio, B., Iniguez, M.A. & Fresno, M. Protein kinase Cζ phosphorylates nuclear factor of activated T cells and regulates its transactivating activity. J. Biol. Chem. 277, 27073–27080 (2002).

    Article  CAS  Google Scholar 

  43. Etienne-Manneville, S. & Hall, A. Cdc42 regulates GSK-3β and adenomatous polyposis coli to control cell polarity. Nature 421, 753–756 (2003).

    Article  CAS  Google Scholar 

  44. Etienne-Manneville, S. et al. Cdc42 and Par6-PKCζ regulate the spatially localized association of Dlg1 and APC to control cell polarization. J. Cell Biol. 170, 895–901 (2005).

    Article  CAS  Google Scholar 

  45. Zhang, J. et al. Antigen receptor-induced activation and cytoskeletal rearrangement are impaired in Wiskott-Aldrich syndrome protein-deficient lymphocytes. J. Exp. Med. 190, 1329–1342 (1999).

    Article  CAS  Google Scholar 

  46. Silvin, C., Belisle, B. & Abo, A. A role for Wiskott-Aldrich syndrome protein in T-cell receptor-mediated transcriptional activation independent of actin polymerization. J. Biol. Chem. 276, 21450–21457 (2001).

    Article  CAS  Google Scholar 

  47. Sommer, K. et al. Phosphorylation of the CARMA1 linker controls NF-κB activation. Immunity 23, 561–574 (2005).

    Article  CAS  Google Scholar 

  48. Thome, M. CARMA1, BCL-10 and MALT1 in lymphocyte development and activation. Nat. Rev. Immunol. 4, 348–359 (2004).

    Article  CAS  Google Scholar 

  49. Badour, K. et al. Fyn and PTP-PEST-mediated regulation of Wiskott-Aldrich syndrome protein (WASp) tyrosine phosphorylation is required for coupling T cell antigen receptor engagement to WASp effector function and T cell activation. J. Exp. Med. 199, 99–112 (2004).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank members of the Miceli lab for critical reading of the manuscript, and J. Ashwell for the antibody to p38 phosphorylated at Tyr323 and for critical reading of the manuscript. The plasmid encoding the NFAT-responsive luciferase reporter construct was a gift from K. Siminovitch (Samuel Lunenfeld Research Institute of Medicine). Supported by the US National Institutes of Health (RO1A1056155 to M.C.M.), the United States Public Health Service (GM07185 to J.L.R.), the University of California at Los Angeles Graduate Division (J.L.R.) and National Institutes of Allergy and Infectious Disease and National Institutes of Health (2-T32-AI-07323 to T.T. and AI07126-30 to L.A.H.). Flow cytometry was done at the UCLA Jonsson Comprehensive Cancer Center and Center for AIDS Research Flow Cytometry Core Facility, which is supported by the National Institutes of Health (CA-16042 and AI-28697).

Author information

Authors and Affiliations

  1. Department of Microbiology, Immunology, and Molecular Genetics, University of California, Los Angeles, 90066, California, USA

    June L Round, Lisa A Humphries, Min Zhang & M Carrie Miceli

  2. Molecular Biology Institute, University of California, Los Angeles, 90066, California, USA

    Tamar Tomassian & M Carrie Miceli

  3. Laboratory of Immune Cell Biology, Center for Cancer Research, National Cancer Institute, Bethesda, 20892, Maryland, USA

    Paul Mittelstadt

Authors
  1. June L Round
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lisa A Humphries
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tamar Tomassian
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Paul Mittelstadt
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Min Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. M Carrie Miceli
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

J.L.R. performed all of the experiments shown; L.A.H provided assistance with calcium flux assays and whole cell lysate p-p38 and anti-phosphotyrosine immunoblots; T.T. provided assistance with mouse work, P.M. provided assistance with phospho-Tyr323 p38 induction and immunoblotting, M.Z. performed anti-phosphotyrosine immunoblots, and M.C.M. supervised all experiments.

Corresponding author

Correspondence to M Carrie Miceli.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

A model whereby MAGUK driven signalosomes coordinate specific activation of NFAT and NF-κB. (PDF 128 kb)

Supplementary Table 1

Primers used to generate GST-Dlgh1 truncations. (PDF 67 kb)

Supplementary Table 2

Primers used to generate Dlgh1 PDZ point mutations. (PDF 67 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Round, J., Humphries, L., Tomassian, T. et al. Scaffold protein Dlgh1 coordinates alternative p38 kinase activation, directing T cell receptor signals toward NFAT but not NF-κB transcription factors. Nat Immunol 8, 154–161 (2007). https://doi.org/10.1038/ni1422

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ni1422

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