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 PKCε–ENH–channel complex specifically modulates N-type Ca2+ channels

Nature Neuroscience volume 6, pages 468–475 (2003)Cite this article

Abstract

Multiple protein kinase C (PKC) isozymes are present in neurons, where they regulate a variety of cellular functions. Due to the lack of specific PKC isozyme inhibitors, it remains unknown how PKC acts on its selective target(s) and achieves its specific actions. Here we show that a PKC binding protein, enigma homolog (ENH), interacts specifically with both PKCε and N-type Ca2+ channels, forming a PKCε–ENH–Ca2+ channel macromolecular complex. Coexpression of ENH facilitated modulation of N-type Ca2+ channel activity by PKC. Disruption of the complex reduced the potentiation of the channel activity by PKC in neurons. Thus, ENH, by interacting specifically with both PKCε and the N-type Ca2+ channel, targets a specific PKC to its substrate to form a functional signaling complex, which is the molecular mechanism for the specificity and efficiency of PKC signaling.

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: Specific interaction between enigma homolog (ENH) and the C-terminus of N-type Ca2+ channels.
Figure 2: Distribution of the ENH protein in the brain.
Figure 3: Cellular localization of ENH in cultured hippocampal neurons.
Figure 4: Formation of a PKCε–ENH–N-type Ca2+ channel complex in neurons.
Figure 5: Facilitation by ENH of PKC modulation of N-type Ca2+ channels.
Figure 6: Responses of chimeric channels to the facilitation of PKC modulation by ENH.
Figure 7: Reduction of the PKC modulation of the N-type Ca2+ current by disruption of the PKCε–ENH–N-type Ca2+ channel complex in neurons.

Similar content being viewed by others

Accession codes

Accessions

GenBank/EMBL/DDBJ

References

  1. Catterall, W.A. Structure and function of neuronal Ca2+ channels and their role in neurotransmitter release. Cell. Calcium 24, 307–323 (1998).

    Article  CAS  Google Scholar 

  2. Levitan, I.B. Modulation of ion channels by protein phosphorylation. How the brain works. Adv. Second Messenger Phosphoprotein Res. 33, 3–22 (1999).

    Article  CAS  Google Scholar 

  3. Vaughan, P.F., Walker, J.H. & Peers, C. The regulation of neurotransmitter secretion by protein kinase C. Mol. Neurobiol. 18, 125–155 (1998).

    Article  CAS  Google Scholar 

  4. Dempsey, E.C. et al. Protein kinase C isozymes and the regulation of diverse cell responses. Am. J. Physiol. 279, 429–438 (2000).

    Article  Google Scholar 

  5. Tanaka, C. & Nishizuka, Y. The protein kinase C family for neuronal signaling. Annu. Rev. Neurosci. 17, 551–567 (1994).

    Article  CAS  Google Scholar 

  6. Dekker, L.V. & Parker, P.J. Protein kinase C: a question of specificity. Trends Biochem. Sci. 19, 73–77 (1994).

    Article  CAS  Google Scholar 

  7. Hofmann, J. The potential for isoenzyme-selective modulation of protein kinase C. FASEB J. 11, 649–669 (1997).

    Article  CAS  Google Scholar 

  8. Way, K.J., Chou, E. & King, G.L. Identification of PKC-isoform-specific biological actions using pharmacological approaches. Trends Pharmacol. Sci. 21, 181–187 (2000).

    Article  CAS  Google Scholar 

  9. Mochly-Rosen, D. Localization of protein kinases by anchoring proteins: a theme in signal transduction. Science 268, 247–251 (1995).

    Article  CAS  Google Scholar 

  10. Pawson, T. & Scott, J.D. Signaling through scaffold, anchoring and adaptor proteins. Science 278, 2075–2080 (1997).

    Article  CAS  Google Scholar 

  11. Mochly-Rosen, D. & Gordon, A.S. Anchoring proteins for protein kinase C: a means for isozyme selectivity. FASEB J. 12, 35–42 (1998).

    Article  CAS  Google Scholar 

  12. Ron, D. & Kazanietz, M.G. New insights into the regulation of protein kinase C and novel phorbol ester receptors. FASEB J. 13, 1658–1676 (1999).

    Article  CAS  Google Scholar 

  13. Zhang, J.F. et al. Distinctive pharmacology and kinetics of cloned neuronal Ca2+ channels and their possible counterparts in mammalian CNS neurons. Neuropharmacology 32, 1075–1088 (1993).

    Article  CAS  Google Scholar 

  14. Dunlap, K., Luebke, J.I. & Turner, T.J. Exocytotic Ca2+ channels in mammalian central neurons. Trends Neurosci. 18, 89–98 (1995).

    Article  CAS  Google Scholar 

  15. Swartz, K.J., Merritt, A., Bean, B.P. & Lovinger, D.M. Protein kinase C modulates glutamate receptor inhibition of Ca2+ channels and synaptic transmission. Nature 361, 165–168 (1993).

    Article  CAS  Google Scholar 

  16. Yang, J. & Tsien, R.W. Enhancement of N- and L-type calcium channel currents by protein kinase C in frog sympathetic neurons. Neuron 10, 127–136 (1993).

    Article  CAS  Google Scholar 

  17. Hell, J.W., Appleyard, S.M., Yokoyama, C.T., Warner, C. & Catterall, W.A. Differential phosphorylation of two size forms of the N-type calcium channel alpha 1 subunit which have different COOH termini. J. Biol. Chem. 269, 7390–7396 (1994).

    CAS  PubMed  Google Scholar 

  18. Stea, A., Soong, T.W. & Snutch, T.P. Determinants of PKC-dependent modulation of a family of neuronal calcium channels. Neuron 15, 929–940 (1995).

    Article  CAS  Google Scholar 

  19. Catterall, W.A. Modulation of sodium and calcium channels by protein phosphorylation and G proteins. Adv. Second Messenger Phosphoprotein Res. 31, 159–181 (1997).

    Article  CAS  Google Scholar 

  20. Malenka, R.C., Madison, D.V. & Nicoll, R.A. Potentiation of synaptic transmission in the hippocampus by phorbol esters. Nature 321, 175–177 (1986).

    Article  CAS  Google Scholar 

  21. Wheeler, D.B., Randall, A. & Tsien, R.W. Roles of N-type and Q-type Ca2+ channels in supporting hippocampal synaptic transmission. Science 264, 107–111 (1994).

    Article  CAS  Google Scholar 

  22. Kuroda, S. et al. Protein-protein interaction of zinc finger LIM domains with protein kinase C. J. Biol. Chem. 271, 31029–31032 (1996).

    Article  CAS  Google Scholar 

  23. Ueki, N. et al. Isolation, tissue expression, and chromosomal assignment of a human LIM protein gene, showing homology to rat enigma homologue (ENH). J. Hum. Genet. 44, 256–260 (1999).

    Article  CAS  Google Scholar 

  24. Dawid, I.B., Breen, J.J. & Toyama, R. LIM domains: multiple roles as adapters and functional modifiers in protein interactions. Trends Genet. 14, 156–162 (1998).

    Article  CAS  Google Scholar 

  25. Bach, I. The LIM domain: regulation by association. Mech. Dev. 91, 5–17 (2000).

    Article  CAS  Google Scholar 

  26. Williams, M.E. et al. Structure and functional expression of an ω-conotoxin-sensitive human N-type calcium channel. Science 257, 389–395 (1992).

    Article  CAS  Google Scholar 

  27. Sudhof, T.C. et al. Synapsins: mosaics of shared and individual domains in a family of synaptic vesicle phosphoproteins. Science 245, 1474–1480 (1989).

    Article  CAS  Google Scholar 

  28. Shapiro, M.S., Zhou, J. & Hille, B. Selective disruption by protein kinases of G-protein-mediated Ca2+ channel modulation. J. Neurophysiol. 76, 311–320 (1996).

    Article  CAS  Google Scholar 

  29. Dodge, F. Jr. & Rahamimoff, R. Co-operative action a calcium ions in transmitter release at the neuromuscular junction. J. Physiol. (Lond.) 193, 419–432 (1967).

    Article  CAS  Google Scholar 

  30. Mintz, I.M., Sabatini, B.L. & Regehr, W.G. Calcium control of transmitter release at a cerebellar synapse. Neuron 15, 675–688 (1995).

    Article  CAS  Google Scholar 

  31. Zhang, J.F., Ellinor, P.T., Aldrich, R.W. & Tsien, R.W. Multiple structural elements in voltage-dependent Ca2+ channels support their inhibition by G proteins. Neuron 17, 991–1003 (1996).

    Article  CAS  Google Scholar 

  32. Johnson, J.A., Gray, M.O., Chen, C.-H. & Mochly-Rosen, D. A protein kinase C translocation inhibitor as an isozyme-selective antagonist of cardiac function. J. Biol. Chem. 271, 24962–24966 (1996).

    Article  CAS  Google Scholar 

  33. Gray, P.C., Scott, J.D. & Catterall, W.A. Regulation of ion channels by cAMP-dependent protein kinase and A-kinase anchoring proteins. Curr. Opin. Neurobiol. 8, 330–334 (1998).

    Article  CAS  Google Scholar 

  34. Fraser, I.D. & Scott, J.D. Modulation of ion channels: a 'current' view of AKAPs. Neuron 23, 423–426 (1999).

    Article  CAS  Google Scholar 

  35. De Waard, M. et al. Direct binding of G-protein betagamma complex to voltage-dependent calcium channels. Nature 385, 446–450 (1997).

    Article  CAS  Google Scholar 

  36. Herlitze, S., Hockerman, G.H., Scheuer, T. & Catterall, W.A. Molecular determinants of inactivation and G protein modulation in the intracellular loop connecting domains I and II of the calcium channel alpha1A subunit. Proc. Natl. Acad. Sci. USA 94, 1512–1516 (1997).

    Article  CAS  Google Scholar 

  37. Qin, N., Platano, D., Olcese, R., Stefani, E. & Birnbaumer, L. Direct interaction of gbetagamma with a C-terminal gbetagamma-binding domain of the Ca2+ channel alpha1 subunit is responsible for channel inhibition by G protein-coupled receptors. Proc. Natl. Acad. Sci. USA 94, 8866–8871 (1997).

    Article  CAS  Google Scholar 

  38. Zamponi, G.W., Bourinet, E., Nelson, D., Nargeot, J. & Snutch, T.P. Crosstalk between G proteins and protein kinase C mediated by the calcium channel alpha1 subunit. Nature 385, 442–446 (1997).

    Article  CAS  Google Scholar 

  39. Lee, A. et al. Ca2+/calmodulin binds to and modulates P/Q-type calcium channels. Nature 399, 155–159 (1999).

    Article  CAS  Google Scholar 

  40. Peterson, B.Z., DeMaria, C.D., Adelman, J.P. & Yue, D.T. Calmodulin is the Ca2+ sensor for Ca2+-dependent inactivation of L-type calcium channels. Neuron 22, 549–558 (1999).

    Article  CAS  Google Scholar 

  41. Zuhlke, R.D., Pitt, G.S., Deisseroth, K., Tsien, R.W. & Reuter, H. Calmodulin supports both inactivation and facilitation of L-type calcium channels. Nature 399, 159–162 (1999).

    Article  CAS  Google Scholar 

  42. Dolphin, A.C. Voltage-dependent calcium channels and their modulation by neurotransmitters and G proteins. Exp. Physiol. 80, 1–36 (1995).

    Article  CAS  Google Scholar 

  43. Klauck, T.M. et al. Coordination of three signaling enzymes by AKAP79, a mammalian scaffold protein. Science 271, 1589–1592 (1996).

    Article  CAS  Google Scholar 

  44. Pawson, T. Protein modules and signalling networks. Nature 373, 573–580 (1995).

    Article  CAS  Google Scholar 

  45. Sheng, M. & Sala, C. PDZ domains and the organization of supremolecular complexes. Annu. Rev. Neurosci. 24, 1–29 (2001).

    Article  CAS  Google Scholar 

  46. Saito, N. et al. Cellular and intracellular localization of epsilon-subspecies of protein kinase C in the rat brain; presynaptic localization of the epsilon-subspecies. Brain Res. 607, 241–248 (1993).

    Article  CAS  Google Scholar 

  47. Khasar, S.G. et al. A novel nociceptor signaling pathway revealed in protein kinase C epsilon mutant mice. Neuron 24, 253–260 (1999).

    Article  CAS  Google Scholar 

  48. Zugaza, J.L., Sinnett-Smith, J., Van Lint, J. & Rozengurt, E. Protein kinase D (PKD) activation in intact cells through a protein kinase C-dependent signal transduction pathway. EMBO J. 15, 6220–6230 (1996).

    Article  Google Scholar 

  49. Zhou, Q., Ruiz-Lozano, P., Martone, M.E. & Chen, J. Cypher, a striated muscle-restricted PDZ and LIM domain-containing protein, binds to alpha-actinin-2 and protein kinase C. J. Biol. Chem. 274, 19807–19813 (1999).

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank R. Pittman, D. Manning, J. Eberwine, J. Meinkoth and C. Deutsch for helpful discussion and critical comments, J. Field for initial help with yeast two-hybrid screening, Z. Lu for oocytes, and M. Maronski and M. Dichter for help with hippocampal neuron cultures. This work was supported by the Penn Research Foundation, American Heart Association, National Institutes of Health (NS39355, J.F.Z.) and a NARSAD young investigator award (Y.M.H.).

Author information

Author notes

  1. Yuka Maeno-Hikichi

    Present address: Department of Cortical Function Disorders, National Institute of Neuroscience, NCNP, 4-1-1 Ogawahigashi, Kodaira, Tokyo, 187-8502, Japan

Authors and Affiliations

  1. Department of Pharmacology, University of Pennsylvania School of Medicine, 3620 Hamilton Walk, Philadelphia, 19104, Pennsylvania, USA

    Yuka Maeno-Hikichi, Shaohua Chang, Meizan Lai, Hong Lin & Ji-fang Zhang

  2. Department of Structural Molecular Biology, Institute of Scientific and Industrial Research, Osaka University, Osaka, 567-0047, Japan

    Kiyoyuki Matsumura, Noritaka Nakagawa & Shun'ichi Kuroda

  3. Institute of Neurological Sciences, University of Pennsylvania School of Medicine, 3620 Hamilton Walk, Philadelphia, 19104, Pennsylvania, USA

    Ji-fang Zhang

Authors
  1. Yuka Maeno-Hikichi
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Shaohua Chang
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Kiyoyuki Matsumura
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Meizan Lai
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Hong Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Noritaka Nakagawa
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Shun'ichi Kuroda
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Ji-fang Zhang
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Ji-fang Zhang.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Maeno-Hikichi, Y., Chang, S., Matsumura, K. et al. A PKCε–ENH–channel complex specifically modulates N-type Ca2+ channels. Nat Neurosci 6, 468–475 (2003). https://doi.org/10.1038/nn1041

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nn1041

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