Mechanism of Auxiliary Subunit Modulation of Neuronal α1E Calcium Channels

Abstract

Voltage-gated calcium channels are composed of a main pore-forming α1 moiety, and one or more auxiliary subunits (β, α2δ) that modulate channel properties. Because modulatory properties may vary greatly with different channels, expression systems, and protocols, it is advantageous to study subunit regulation with a uniform experimental strategy. Here, in HEK 293 cells, we examine the expression and activation gating of α1E calcium channels in combination with a β (β1–β4) and/or the α2δ subunit, exploiting both ionic- and gating-current measurements. Furthermore, to explore whether more than one auxiliary subunit can concomitantly specify gating properties, we investigate the effects of cotransfecting α2δ with β subunits, of transfecting two different β subunits simultaneously, and of COOH-terminal truncation of α1E to remove a second β binding site. The main results are as follows. (a) The α2δ and β subunits modulate α1E in fundamentally different ways. The sole effect of α2δ is to increase current density by elevating channel density. By contrast, though β subunits also increase functional channel number, they also enhance maximum open probability (Gmax/Qmax) and hyperpolarize the voltage dependence of ionic-current activation and gating-charge movement, all without discernible effect on activation kinetics. Different β isoforms produce nearly indistinguishable effects on activation. However, β subunits produced clear, isoform-specific effects on inactivation properties. (b) All the β subunit effects can be explained by a gating model in which subunits act only on weakly voltage-dependent steps near the open state. (c) We find no clear evidence for simultaneous modulation by two different β subunits. (d) The modulatory features found here for α1E do not generalize uniformly to other α1 channel types, as α1C activation gating shows marked β isoform dependence that is absent for α1E. Together, these results help to establish a more comprehensive picture of auxiliary-subunit regulation of α1E calcium channels.

The N-terminal Domain Tethers the Voltage-gated Calcium Channel β2e-subunit to the Plasma Membrane via Electrostatic and Hydrophobic Interactions

“…8D). Half-inactivation voltages (V 0.5 ) obtained from the fit to a Boltzmann distribution for Ca V 2.3 coexpressed with ␤ 2e WT and mutant were similar to those previously reported for membrane-anchored ␤ 2a and the cytosolic ␤ 1b , respectively (12).…”

Diversity and Developmental Expression of L-type Calcium Channel β2 Proteins and Their Influence on Calcium Current in Murine Heart

“…Steady-state activation and inactivation has been shown previously to be affected by the type of coexpressed CaV␤ subunit in heterologous expression studies (37,38). As shown in Fig.…”

Alternative Splicing of the β4Subunit Has α1Subunit Subtype-Specific Effects on Ca2+Channel Gating

“…Their studies yielded five results pertinent to our findings. (1) Relative to ␣ 1E alone, all ␤ subunit constructs tested caused a nearly identical hyperpolarizing shift in the V 1/2 of activation and decrease in slope factor [see also Jones et al (1998)]. (2) Deletion of the N terminus of the ␤ 1b , ␤ 2a , and ␤ 3 subunits had no effect on the fast component of activation.…”