Characterization of Ca v 1.4 complexes (α 1 1.4, β 2 , and α 2 δ 4 ) in HEK293T cells and in the retina

Abstract

In photoreceptor synaptic terminals, voltage-gated Ca v 1.4 channels mediate Ca 2+ signals required for transmission of visual stimuli. Like other high voltage-activated Ca v channels, Ca v 1.4 channels are composed of a main pore-forming Ca v 1.4 α 1 subunit and auxiliary β and α 2 δ subunits. Of the four distinct classes of β and α 2 δ, β 2 and α 2 δ 4 are thought to co-assemble with Ca v 1.4 α 1 subunits in photoreceptors. However, an understanding of the functional properties of this combination of Ca v subunits is lacking. Here, we provide evidence that Ca v 1.4 α 1 , β 2 , and α 2 δ 4 contribute to Ca v 1.4 channel complexes in the retina and describe their properties in electrophysiological recordings. In addition, we identified a variant of β 2 , named here β 2X13 , which, along with β 2a , is present in photoreceptor terminals. Ca v 1.4 α 1 , β 2 , and α 2 δ 4 were coimmunoprecipitated from lysates of transfected HEK293 cells and mouse retina and were found to interact in the outer plexiform layer of the retina containing the photoreceptor synaptic terminals, by proximity ligation assays. In whole-cell patch clamp recordings of transfected HEK293T cells, channels (Ca v 1.4 α 1 + β 2X13 ) containing α 2 δ 4 exhibited weaker voltage-dependent activation than those with α 2 δ 1 . Moreover, compared with channels (Ca v 1.4 α 1 + α 2 δ 4 ) with β 2a , β 2X13 -containing channels exhibited greater voltage-dependent inactivation. The latter effect was specific to Ca v 1.4 because it was not seen for Ca v 1.2 channels. Our results provide the first detailed functional analysis of the Ca v 1.4 subunits that form native photoreceptor Ca v 1.4 channels and indicate potential heterogeneity in these channels conferred by β 2a and β 2X13 variants.