Structural determinants of CaV1.3 L-type calcium channel gating

A C-terminal modulatory domain (CTM) tightly regulates the biophysical properties of Ca(v)1.3 L-type Ca(2+) channels, in particular the voltage dependence of activation (V(0.5)) and Ca(2+) dependent inactivation (CDI). A functional CTM is present in the long C-terminus of human and mouse Ca(v)1.3 (Ca(v)1.3(L)), but not in a rat long cDNA clone isolated from superior cervical ganglia neurons (rCa(v)1.3(scg)). We therefore addressed the question if this represents a species-difference and compared the biophysical properties of rCa(v)1.3(scg) with a rat cDNA isolated from rat pancreas (rCa(v)1.3(L)). When expressed in tsA-201 cells under identical experimental conditions rCa(v)1.3(L) exhibited Ca(2+) current properties indistinguishable from human and mouse Ca(v)1.3(L), compatible with the presence of a functional CTM. In contrast, rCa(v)1.3(scg) showed gating properties similar to human short splice variants lacking a CTM. rCa(v)1.3(scg) differs from rCa(v)1.3(L) at three single amino acid (aa) positions, one alternative spliced exon (exon31), and a N-terminal polymethionine stretch with two additional lysines. Two aa (S244, A2075) in rCa(v)1.3(scg) explained most of the functional differences to rCa(v)1.3(L). Their mutation to the corresponding residues in rCa(v)1.3(L) (G244, V2075) revealed that both contributed to the more negative V 0.5, but caused opposite effects on CDI. A2075 (located within a region forming the CTM) additionally permitted higher channel open probability. The cooperative action in the double-mutant restored gating properties similar to rCa(v)1.3(L). We found no evidence for transcripts containing one of the single rCa(v)1.3(scg) mutations in rat superior cervical ganglion preparations. However, the rCa(v)1.3(scg) variant provided interesting insight into the structural machinery involved in Ca(v)1.3 gating.