Mechanism of auxiliary subunit modulation of neuronal α1E calcium channels

Voltage-gated calcium channels are composed of a main pore-forming oil moiety, and one or more auxiliary subunits (beta, alpha(2)delta) 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. Were, in HEK 293 cells, we examine the expression and activation gating of alpha(1E) calcium channels in combination with a beta (beta(1)-beta(4)) and/or the alpha(2)delta 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 alpha(2)delta with beta subunits, of transfecting two different beta subunits simultaneously, and of COOH-terminal truncation of alpha(1E) to remove a second beta binding site. The main results are as follows. (a) The alpha(2)delta and beta subunits modulate alpha(1E) in fundamentally different ways. The sole effect of alpha(2)delta is to increase current density by elevating channel density. By contrast, though beta subunits also increase functional channel number, they also enhance maximum open probability (G(max)/Q(max)) and hyperpolarize the voltage dependence of ionic-current activation and gating-charge movement, all without discernible effect on activation kinetics. Different beta isoforms produce nearly indistinguishable effects on activation. However, beta subunits produced clear, isoform-specific effects on inactivation properties. (b) All the beta 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 beta subunits. (d) The modulatory features found here for alpha(1E) do not generalize uniformly to other oil channel types, as cc,, activation gating shows marked beta isoform dependence that is absent for alpha(1E) Together, these results help to establish a more comprehensive picture of auxiliary-subunit regulation of alpha(1E) calcium channels.