Abstract
The β-subunit of high-voltage-activated (HVA) calcium channels is essential for the regulation of expression and gating. On the other hand, various reports have suggested that β subunits play no role in the regulation of low-voltage-activated T-type channels. In addition there has been no clear demonstration of a physical interaction between the α-subunit of T-type channel with β-subunit. In this study, we systematically investigated the interaction between CaVα and CaVβ. The four CaVβ isoforms were expressed in a bacterial system and purified into homogeneity, whereas the ten types of CaVα alpha interaction domain (AID) peptides were chemically synthesized. All possible combinations of CaVα and CaVβ were then tested for by in vitro immunoassays. We describe here the identification of a new interaction between CaV3.3 and CaVβ proteins. This interaction is of low affinity compared to that between the AID of the HVA α-subunit and the alpha-binding pocket (ABP) site of the β-subunit. The AID peptide of HVA channel exerted no effect on the CaV3.3-CaVβ interaction, thus demonstrating that another site not in the ABP of CaVβ protein played a role in binding with CaV3.3. This is the first demonstration of an α-β subunit interaction in a T-type calcium channel.
Similar content being viewed by others
References
Arias, J.M., Murbartian, J., Vitko, I., Lee, J.H., and Perez-Reyes, E. (2005). Transfer of b subunit regulation from high to low voltagegated Ca2+ channels. FEBS Lett. 579, 3907–3912.
Arikkath, J., and Campbell, K.P. (2003). Auxiliary subunits: essential components of the voltage-gated calcium channel complex. Curr. Opin. Neurobiol. 13, 298–307.
Ball, S.L., Powers, P.A., Shin, H.S., Morgans, C.W., Peachey, N.S., and Gregg, R.G. (2002). Role of the β2 subunit of voltage-dependent calcium channels in the retinal outer plexiform layer. Invest. Ophthalmol. Vis. Sci. 43, 1595–1603.
Bean, B.P. (1989). Classes of calcium channels in vertebrate cells. Annu. Rev. Physiol. 51, 367–384.
Bourgeas, R., Basse, M.J., Morelli, X., and Roche, P. (2010). Atomic analysis of protein-protein interfaces with known inhibitors: the 2P2I database. PLoS One 5, e9598.
Calakos, N., Bennett, M.K., Peterson, K.E., and Scheller, R.H. (1994). Protein-protein interactions contributing to the specificity of intracellular vesicular trafficking. Science 263, 1146–1149.
Canti, C., Davies, A., Berrow, N.S., Butcher, A.J., Page, K.M., and Dolphin, A.C. (2001). Evidence for two concentration-dependent processes for beta-subunit effects on alpha1B calcium channels. Biophys. J. 81, 1439–1451.
Catterall, W.A. (2000). Structure and regulation of voltage-gated Ca2+ channels. Annu. Rev. Cell Dev. Biol. 16, 521–555.
Chen, C.C., Lamping, K.G., Nuno, D.W., Barresi, R., Prouty, S.J., Lavoie, J.L., Cribbs, L.L., England, S.K., Sigmund, C.D., Weiss, R.M., et al. (2003). Abnormal coronary function in mice deficient in α1H T-type Ca2+ channels. Science 302, 1416–1418.
De Waard, M., Witcher, D.R., Pragnell, M., Liu, H., and Campbell, K.P. (1995). Properties of the α1b anchoring site in voltage-dependent Ca2+ channels. J. Biol. Chem. 270, 12056–12064.
Dolphin, A.C., Wyatt, C.N., Richards, J., Beattie, R.E., Craig, P., Lee, J.H., Cribbs, L.L., Volsen, S.G., and Perez-Reyes, E. (1999). The effect of a2d and other accessory subunits on expression and properties of the calcium channel α1G. J. Physiol. 519, 35–45.
Dubel, S.J., Altier, C., Chaumont, S., Lory, P., Bourinet, E., and Nargeot, J. (2004). Plasma membrane expression of T-type calcium channel α1 subunits is modulated by high voltage-activated auxiliary subunits. J. Biol. Chem. 279, 29263–29269.
Ellinor, P.T., Zhang, J.F., Randall, A.D., Zhou, M., Schwarz, T.L., Tsien, R.W., and Horne, W.A. (1993). Functional expression of a rapidly inactivating neuronal calcium channel. Nature 363, 455–458.
Geib, S., Sandoz, G., Mabrouk, K., Matavel, A., Marchot, P., Hoshi, T., Villaz, M., Ronjat, M., Miquelis, R., Leveque, C., et al. (2002). Use of a purified and functional recombinant calcium-channel β4 subunit in surface-plasmon resonance studies. Biochem. J. 364, 285–292.
Hanlon, M.R., Berrow, N.S., Dolphin, A.C., and Wallace, B.A. (1999). Modelling of a voltage-dependent Ca2+ channel beta subunit as a basis for understanding its functional properties. FEBS Lett. 445, 366–370.
Huguenard, J.R. (1996). Low-threshold calcium currents in central nervous system neurons. Annu. Rev. Physiol. 58, 329–348.
Jun, K., Piedras-Renteria, E.S., Smith, S.M., Wheeler, D.B., Lee, S.B., Lee, T.G., Chin, H., Adams, M.E., Scheller, R.H., Tsien, R.W., et al. (1999). Ablation of P/Q-type Ca2+ channel currents, altered synaptic transmission, and progressive ataxia in mice lacking the α1A-subunit. Proc. Natl. Acad. Sci. USA 96, 15245–15250.
Lacerda, A.E., Perez-Reyes, E., Wei, X., Castellano, A., and Brown, A.M. (1994). T-type and N-type calcium channels of Xenopus oocytes: evidence for specific interactions with β subunits. Biophys. J. 66, 1833–1843.
Lambert, R.C., Maulet, Y., Mouton, J., Beattie, R., Volsen, S., De Waard, M., and Feltz, A. (1997). T-type Ca2+ current properties are not modified by Ca2+ channel β subunit depletion in nodosus ganglion neurons. J. Neurosci. 17, 6621–6628.
Leuranguer, V., Bourinet, E., Lory, P., and Nargeot, J. (1998). Antisense depletion of β-subunits fails to affect T-type calcium channels properties in a neuroblastoma cell line. Neuropharmacology 37, 701–708.
Ludwig, A., Flockerzi, V., and Hofmann, F. (1997). Regional expression and cellular localization of the α1 and β subunit of high voltage-activated calcium channels in rat brain. J. Neurosci. 17, 1339–1349.
Mikami, A., Imoto, K., Tanabe, T., Niidome, T., Mori, Y., Takeshima, H., Narumiya, S., and Numa, S. (1989) Primary structure and functional expression of the cardiac dihydropyridine-sensitive calcium channel. Nature 340, 230–233.
Monteil, A., Chemin, J., Leuranguer, V., Altier, C., Mennessier, G., Bourinet, E., Lory, P., and Nargeot, J. (2000). Specific properties of T-type calcium channels generated by the human α1I subunit. J. Biol. Chem. 275, 16530–16535.
Na, H.S., Choi, S., Kim, J., Park, J., and Shin, H.S. (2008). Attenuated neuropathic pain in CaV3.1 null mice. Mol. Cells 25, 242–246.
Nowycky, M.C., Fox, A.P., and Tsien, R.W. (1985). Three types of neuronal calcium channel with different calcium agonist sensitivity. Nature 316, 440–443.
Opatowsky, Y., Chomsky-Hecht, O., Kang, M.G., Campbell, K.P., and Hirsch, J.A. (2003). The voltage-dependent calcium channel beta subunit contains two stable interacting domains. J. Biol. Chem. 278, 52323–52332.
Pawson, T., and Nash, P. (2003). Assembly of cell regulatory systems through protein interaction domains. Science 300, 445–452.
Perez-Reyes, E. (2006). Molecular characterization of T-type calcium channels. Cell Calcium 40, 89–96.
Perez-Reyes, E., Cribbs, L.L., Daud, A., Lacerda, A.E., Barclay, J., Williamson, M.P., Fox, M., Rees, M., and Lee, J.H. (1998). Molecular characterization of a neuronal low-voltage-activated Ttype calcium channel. Nature 391, 896–900.
Qin, N., Platano, D., Olcese, R., Stefani, E., and Birnbaumer, L. (1997). Direct interaction of Gβγ with a C-terminal Gβγ-binding domain of the Ca2+ channel α1 subunit is responsible for channel inhibition by G protein-coupled receptors. Proc. Natl. Acad. Sci. USA 94, 8866–8871.
Richards, M.W., Butcher, A.J., and Dolphin, A.C. (2004) Ca2+ channel β-subunits: structural insights AID our understanding. Trends Pharmacol. Sci. 25, 626–632.
Talley, E.M., Cribbs, L.L., Lee, J.H., Daud, A., Perez-Reyes, E., and Bayliss, D.A. (1999). Differential distribution of three members of a gene family encoding low voltage-activated (T-type) calcium channels. J. Neurosci. 19, 1895–1911.
Van Petegem, F., Clark, K.A., Chatelain, F.C., and Minor, D.L., Jr. (2004). Structure of a complex between a voltage-gated calcium channel β-subunit and an α-subunit domain. Nature 429, 671–675.
Walker, D., Bichet, D., Campbell, K.P., and De Waard, M. (1998). A β4 isoform-specific interaction site in the carboxyl-terminal region of the voltage-dependent Ca2+ channel α1A subunit. J. Biol. Chem. 273, 2361–2367.
Author information
Authors and Affiliations
Corresponding author
About this article
Cite this article
Bae, J., Suh, E.J. & Lee, C. Interaction of T-type calcium channel CaV3.3 with the β-subunit. Mol Cells 30, 185–191 (2010). https://doi.org/10.1007/s10059-010-0106-z
Received:
Revised:
Accepted:
Published:
Issue Date:
DOI: https://doi.org/10.1007/s10059-010-0106-z