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

Abstract

Coexpression of the cloned voltage-dependent Ca 2+channel α 2 δ subunit with the pore-forming α 1 subunit 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 α 2 δ complex or the δ subunit on the Ca 2+channel α 1 subunit. Transient transfection of tsA201 cells with the cardiac L-type α 1C subunit alone resulted in the expression of inward voltage-activated currents as well as measurable [3 H]-PN200-110 binding to membranes from transfected cells. Coexpression of the α 2 δ subunit significantly increased the macroscopic current amplitude, altered the voltage dependence and the kinetics of the current, and enhanced [3 H]-PN200-110 binding. Except for the increase in amplitude, coexpression of the δ subunit reproduced entirely the effects of the full-length α 2 δ subunit on the biophysical properties of the α 1C currents. However, no effect on specific [3 H]-PN200-110 binding was observed on δ subunit coexpression. Likewise, profound effects on current kinetics of the neuronal α 1A subunit were observed on coexpression of the α 2 δ complex in Xenopus oocytes. 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 α 2 δ regulation are conserved across L-type and non-L type Ca 2+channels. Taken together, our results indicate that the region of the α 2 δ 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 α 2 domain of the α 2 δ complex.