The R-Domain: Identification of an N-terminal Region of the α2δ-1 Subunit Which is Necessary and Sufficient for its Effects on Cav2.2 Calcium Currents

Voltage-gated calcium channels (Cav) and their associated proteins are pivotal signalling complexes in excitable cell physiology. In nerves and muscle, Cav tailor calcium influx to processes including neurotransmission, muscle contraction and gene expression. Cav comprise a pore-forming α1 and modulatory β and α2δ subunits – the latter targeted by anti-epileptic and anti-nociceptive gabapentinoid drugs. However, the mechanisms of gabapentinoid action are unclear, not least because detailed structure-function mapping of the α2δ subunit remains lacking. Using molecular biology and electrophysiological approaches we have conducted the first systematic mapping of α2δ subunit structurefunction. We generated a series of cDNA constructs encoding chimera, from which successive amino acids from the rat α2δ-1 subunit were incorporated into a Type 1 reporter protein – PIN-G, to produce sequential extensions from the transmembrane (TM) region towards the N-terminus. By successive insertion of a TGA stop codon, a further series of N- to Cterminal extension constructs lacking the TM region, were also generated. Using this approach we have defined the minimal region of α2δ-1 - we term the R-domain (Rd), that appears to contain all the machinery necessary to support the electrophysiological and trafficking effects of α2δ-1 on Cav. Structural algorithms predict that Rd is conserved across all four α2δ subunits, including RNA splice variants, and irrespective of phyla and taxa. We suggest, therefore, that Rd likely constitutes the major locus for physical interaction with the α1 subunit and may provide a target for novel Cav therapeutics.