Dissection of Functional Domains of the Voltage-Dependent Ca2+Channel α2δ Subunit

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Abstract
Coexpression of the cloned voltage-dependent Ca2+channel α2δ subunit with the pore-forming α1subunit results in a significant increase in macroscopic current amplitude. To gain insight into the mechanism underlying this interaction, we have examined the regulatory effect of either the α2δ complex or the δ subunit on the Ca2+channel α1subunit. Transient transfection of tsA201 cells with the cardiac L-type α1Csubunit alone resulted in the expression of inward voltage-activated currents as well as measurable [3H]-PN200-110 binding to membranes from transfected cells. Coexpression of the α2δ subunit significantly increased the macroscopic current amplitude, altered the voltage dependence and the kinetics of the current, and enhanced [3H]-PN200-110 binding. Except for the increase in amplitude, coexpression of the δ subunit reproduced entirely the effects of the full-length α2δ subunit on the biophysical properties of the α1Ccurrents. However, no effect on specific [3H]-PN200-110 binding was observed on δ subunit coexpression. Likewise, profound effects on current kinetics of the neuronal α1Asubunit were observed on coexpression of the α2δ complex inXenopusoocytes. Furthermore, by using a chimeric strategy, we localized the region involved in this regulation to the transmembrane domain of the δ subunit. These data strongly suggest that the molecular determinants involved in α2δ regulation are conserved across L-type and non-L type Ca2+channels. Taken together, our results indicate that the region of the α2δ subunit involved in the modulation of the gating properties of the high voltage-activated calcium channels is localized in the δ domain of the protein. In contrast, the level of membrane expression of functional channels relies on the presence of the α2domain of the α2δ complex.

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