The α2/δ subunit of voltage sensitive Ca²⁺ channels is a single transmembrane extracellular protein which is involved in regulated secretion

Abstract

The membrane topology of α2/δ subunit was investigated utilizing electrophysiological functional assay and specific anti‐α2 antibodies. (a) cRNA encoding a deleted α2/δ subunit was coinjected with α1C subunit of the L‐type calcium channel into Xenopus oocytes. The truncated form, lacking the third putative TM domain (α2/δΔTMIII), failed to amplify the expressed inward currents, normally induced by α 1c coinjected with intact α2/δ subunit. Western blot analysis of α2/δΔTMIII shows the appearance of a degraded α2 protein and no expression of the full‐size two‐TM truncated‐protein. The improper processing of α2/δΔTMIII suggests that the α2/δ is a single TM domain protein and the TM region is positioned at the δ subunit. (b) External application of anti‐α2 antibodies, prepared for an epitope within the alternatively spliced and ‘intracellular’ region, inhibits depolarization induced secretion in PC12, further supporting an external location of the α2 subunit and establishing δ subunit as the only membrane anchor for the extracellular α2 subunit.