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:

A voltage-gated sodium channel is essential for the positive selection of CD4+ T cells

Abstract

The sustained entry of Ca2+ into CD4+CD8+ double-positive thymocytes is required for positive selection. Here we identified a voltage-gated Na+ channel (VGSC) that was essential for positive selection of CD4+ T cells. Pharmacological inhibition of VGSC activity inhibited the sustained Ca2+ influx induced by positively selecting ligands and the in vitro positive selection of CD4+ but not CD8+ T cells. In vivo short hairpin RNA (shRNA)-mediated knockdown of the gene encoding a regulatory β-subunit of a VGSC specifically inhibited the positive selection of CD4+ T cells. Ectopic expression of VGSC in peripheral AND CD4+ T cells bestowed the ability to respond to a positively selecting ligand, which directly demonstrated that VGSC expression was responsible for the enhanced sensitivity. Thus, active VGSCs in thymocytes provide a mechanism by which a weak positive selection signal can induce the sustained Ca2+ signals required for CD4+ T cell development.

This is a preview of subscription content, access via your institution

Access options

Rent or buy this article

Prices vary by article type

from$1.95

to$39.95

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Sustained Ca2+ signals in AND DP thymocytes induced by gp250 ligand.
Figure 2: Expression of Scn5a and Scn4b during the course of positive selection.
Figure 3: The pore-forming SCN5A subunit is critical in positive selection.
Figure 4: A regulatory subunit SCN4B is critical for positive selection.
Figure 5: Knockdown of Scn5a by shRNA impairs the positive selection of CD4SP cells in vivo.
Figure 6: Peripheral AND CD4+ T cells acquire the ability to respond to positively selecting ligands by expression of human VGSCs.

Similar content being viewed by others

Accession codes

Primary accessions

Gene Expression Omnibus

References

  1. Starr, T.K., Jameson, S.C. & Hogquist, K.A. Positive and negative selection of T cells. Annu. Rev. Immunol. 21, 139–176 (2003).

    Article  CAS  Google Scholar 

  2. Moran, A.E. & Hogquist, K.A. T-cell receptor affinity in thymic development. Immunology 135, 261–267 (2012).

    Article  CAS  Google Scholar 

  3. Morris, G.P. & Allen, P.M. How the TCR balances sensitivity and specificity for the recognition of self and pathogens. Nat. Immunol. 13, 121–128 (2012).

    Article  CAS  Google Scholar 

  4. Kane, L.P. & Hedrick, S.M. A role for calcium influx in setting the threshold for CD4+CD8+ thymocyte negative selection. J. Immunol. 156, 4594–4601 (1996).

    CAS  Google Scholar 

  5. Mariathasan, S. et al. Duration and strength of extracellular signal-regulated kinase signals are altered during positive versus negative thymocyte selection. J. Immunol. 167, 4966–4973 (2001).

    Article  CAS  Google Scholar 

  6. Bhakta, N.R., Oh, D.Y. & Lewis, R.S. Calcium oscillations regulate thymocyte motility during positive selection in the three-dimensional thymic environment. Nat. Immunol. 6, 143–151 (2005).

    Article  CAS  Google Scholar 

  7. Daniels, M.A. et al. Thymic selection threshold defined by compartmentalization of Ras/MAPK signalling. Nature 444, 724–729 (2006).

    Article  CAS  Google Scholar 

  8. Oh-hora, M. Calcium signaling in the development and function of T-lineage cells. Immunol. Rev. 231, 210–224 (2009).

    Article  CAS  Google Scholar 

  9. Werlen, G., Hausmann, B. & Palmer, E. A motif in the alphabeta T-cell receptor controls positive selection by modulating ERK activity. Nature 406, 422–426 (2000).

    Article  CAS  Google Scholar 

  10. McNeil, L.K., Starr, T.K. & Hogquist, K.A. A requirement for sustained ERK signaling during thymocyte positive selection in vivo. Proc. Natl. Acad. Sci. USA 102, 13574–13579 (2005).

    Article  CAS  Google Scholar 

  11. Cahalan, M.D. & Chandy, K.G. The functional network of ion channels in T lymphocytes. Immunol. Rev. 231, 59–87 (2009).

    Article  CAS  Google Scholar 

  12. Feske, S. ORAI1 and STIM1 deficiency in human and mice: roles of store-operated Ca2+ entry in the immune system and beyond. Immunol. Rev. 231, 189–209 (2009).

    Article  CAS  Google Scholar 

  13. Gwack, Y. et al. Hair loss and defective T- and B-cell function in mice lacking ORAI1. Mol. Cell. Biol. 28, 5209–5222 (2008).

    Article  CAS  Google Scholar 

  14. Vig, M. et al. Defective mast cell effector functions in mice lacking the CRACM1 pore subunit of store-operated calcium release-activated calcium channels. Nat. Immunol. 9, 89–96 (2008).

    Article  CAS  Google Scholar 

  15. Beyersdorf, N. et al. STIM1-independent T cell development and effector function in vivo. J. Immunol. 182, 3390–3397 (2009).

    Article  CAS  Google Scholar 

  16. Lo, W.L. et al. An endogenous peptide positively selects and augments the activation and survival of peripheral CD4+ T cells. Nat. Immunol. 10, 1155–1161 (2009).

    Article  CAS  Google Scholar 

  17. Yu, F.H. et al. Sodium channel beta4, a new disulfide-linked auxiliary subunit with similarity to beta2. J. Neurosci. 23, 7577–7585 (2003).

    Article  CAS  Google Scholar 

  18. Grieco, T.M., Malhotra, J.D., Chen, C., Isom, L.L. & Raman, I.M. Open-channel block by the cytoplasmic tail of sodium channel beta4 as a mechanism for resurgent sodium current. Neuron 45, 233–244 (2005).

    Article  CAS  Google Scholar 

  19. Bant, J.S. & Raman, I.M. Control of transient, resurgent, and persistent current by open-channel block by Na channel beta4 in cultured cerebellar granule neurons. Proc. Natl. Acad. Sci. USA 107, 12357–12362 (2010).

    Article  CAS  Google Scholar 

  20. Catterall, W.A. From ionic currents to molecular mechanisms: the structure and function of voltage-gated sodium channels. Neuron 26, 13–25 (2000).

    Article  CAS  Google Scholar 

  21. Goldin, A.L. et al. Nomenclature of voltage-gated sodium channels. Neuron 28, 365–368 (2000).

    Article  CAS  Google Scholar 

  22. Fozzard, H.A. & Lipkind, G.M. The tetrodotoxin binding site is within the outer vestibule of the sodium channel. Mar. Drugs 8, 219–234 (2010).

    Article  CAS  Google Scholar 

  23. Kaczmarek, L.K. Non-conducting functions of voltage-gated ion channels. Nat. Rev. Neurosci. 7, 761–771 (2006).

    Article  CAS  Google Scholar 

  24. Brackenbury, W.J., Djamgoz, M.B. & Isom, L.L. An emerging role for voltage-gated Na+ channels in cellular migration: regulation of central nervous system development and potentiation of invasive cancers. Neuroscientist 14, 571–583 (2008).

    Article  CAS  Google Scholar 

  25. Davey, G.M. et al. Preselection thymocytes are more sensitive to T cell receptor stimulation than mature T cells. J. Exp. Med. 188, 1867–1874 (1998).

    Article  CAS  Google Scholar 

  26. Painter, M.W., Davis, S., Hardy, R.R., Mathis, D. & Benoist, C. Transcriptomes of the B and T lineages compared by multiplatform microarray profiling. J. Immunol. 186, 3047–3057 (2011).

    Article  CAS  Google Scholar 

  27. Catterall, W.A. Structure and regulation of voltage-gated Ca2+ channels. Annu. Rev. Cell Dev. Biol. 16, 521–555 (2000).

    Article  CAS  Google Scholar 

  28. Dravid, S.M., Baden, D.G. & Murray, T.F. Brevetoxin activation of voltage-gated sodium channels regulates Ca dynamics and ERK1/2 phosphorylation in murine neocortical neurons. J. Neurochem. 89, 739–749 (2004).

    Article  CAS  Google Scholar 

  29. Fekete, A. et al. Mechanism of the persistent sodium current activator veratridine-evoked Ca elevation: implication for epilepsy. J. Neurochem. 111, 745–756 (2009).

    Article  CAS  Google Scholar 

  30. Catterall, W.A. Signaling complexes of voltage-gated sodium and calcium channels. Neurosci. Lett. 486, 107–116 (2010).

    Article  CAS  Google Scholar 

  31. Rook, M.B., Evers, M.M., Vos, M.A. & Bierhuizen, M.F. Biology of cardiac sodium channel Nav1.5 expression. Cardiovasc. Res. 93, 12–23 (2012).

    Article  CAS  Google Scholar 

  32. Omilusik, K. et al. The Ca(v)1.4 calcium channel is a critical regulator of T cell receptor signaling and naive T cell homeostasis. Immunity 35, 349–360 (2011).

    Article  CAS  Google Scholar 

  33. Juang, J. et al. Peptide-MHC heterodimers show that thymic positive selection requires a more restricted set of self-peptides than negative selection. J. Exp. Med. 207, 1223–1234 (2010).

    Article  CAS  Google Scholar 

  34. Singer, A. Molecular and cellular basis of T cell lineage commitment: An overview. Semin. Immunol. 22, 253 (2010).

    Article  Google Scholar 

  35. Papadatos, G.A. et al. Slowed conduction and ventricular tachycardia after targeted disruption of the cardiac sodium channel gene Scn5a. Proc. Natl. Acad. Sci. USA 99, 6210–6215 (2002).

    Article  CAS  Google Scholar 

  36. Allen, P.M. Themis imposes new law and order on positive selection. Nat. Immunol. 10, 805–806 (2009).

    Article  CAS  Google Scholar 

  37. Sun, G. et al. The zinc finger protein cKrox directs CD4 lineage differentiation during intrathymic T cell positive selection. Nat. Immunol. 6, 373–381 (2005).

    Article  CAS  Google Scholar 

  38. He, X. et al. The zinc finger transcription factor Th-POK regulates CD4 versus CD8 T-cell lineage commitment. Nature 433, 826–833 (2005).

    Article  CAS  Google Scholar 

  39. Gallo, E.M. et al. Calcineurin sets the bandwidth for discrimination of signals during thymocyte development. Nature 450, 731–735 (2007).

    Article  CAS  Google Scholar 

  40. Albu, D.I. et al. BCL11B is required for positive selection and survival of double-positive thymocytes. J. Exp. Med. 204, 3003–3015 (2007).

    Article  CAS  Google Scholar 

  41. Clements, C.S. et al. The production, purification and crystallization of a soluble heterodimeric form of a highly selected T-cell receptor in its unliganded and liganded state. Acta Crystallogr. D Biol. Crystallogr. 58, 2131–2134 (2002).

    Article  Google Scholar 

  42. Weber, K.S., Hildner, K., Murphy, K.M. & Allen, P.M. Trpm4 differentially regulates Th1 and Th2 function by altering calcium signaling and NFAT localization. J. Immunol. 185, 2836–2846 (2010).

    Article  CAS  Google Scholar 

  43. Fanger, C.M., Neben, A.L. & Cahalan, M.D. Differential Ca2+ influx, KCa channel activity, and Ca2+ clearance distinguish Th1 and Th2 lymphocytes. J. Immunol. 164, 1153–1160 (2000).

    Article  CAS  Google Scholar 

  44. Allen, P.M. Construction of murine T-T-cell hybridomas. in Monoclonal Antibody Production Techniques and Applications. (Marcel Dekker Inc., New York, 1987).

Download references

Acknowledgements

We thank A. Farr (University of Washington) for the ANV41.2 the cortical epithelial cell line; D. Kreamalmeyer for maintaining the mouse colony; S. Horvath for peptide synthesis and purification; M. Cella for advice on the production of the SCN4B-immunoglobulin fusion protein; M. Colonna (Washington University School of Medicine) for the Ceacam4-Ig fusion protein; S. Chan, S. Srivatsan and D. Bhattacharya for advice on shRNA-mediated knockdown experiments; R. Schreiber for mouse IFN-γ; J. Silva, C. Nichols and S. Goldstein (University of Chicago) for the human SCN5AGFP construct; J. Nerbonne for discussions; N. Felix for initiating the gp250 project; G. Morris for critical data analysis and assistance with the manuscript; and K. Weber, C. Morley, A. Shaw, M. Vig, Y. Huang, K. Murphy and T. Egawa for critical reading of the manuscript and comments. Supported by the US National Institutes of Health (AI-24157 to P.M.A.).

Author information

Authors and Affiliations

Authors

Contributions

W.-L.L. and P.M.A. designed the study and wrote the manuscript; W.-L.L. did the experimental work; and D.L.D. generated the AND hybridoma cell lines.

Corresponding author

Correspondence to Paul M Allen.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–5 (PDF 754 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Lo, WL., Donermeyer, D. & Allen, P. A voltage-gated sodium channel is essential for the positive selection of CD4+ T cells. Nat Immunol 13, 880–887 (2012). https://doi.org/10.1038/ni.2379

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ni.2379

This article is cited by

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