Alternative splicing at C terminus of Ca v1.4 calcium channel modulates calcium-dependent inactivation, activation potential, and current density

Abstract

The Ca V1.4 voltage-gated calcium channel is predominantly expressed in the retina, and mutations to this channel have been associated with human congenital stationary night blindness type-2. The L-type Ca V1.4 channel displays distinct properties such as absence of calcium-dependent inactivation (CDI) and slow voltage-dependent inactivation (VDI) due to the presence of an autoinhibitory domain (inhibitor of CDI) in the distal C terminus. We hypothesized that native Ca V1.4 is subjected to extensive alternative splicing, much like the other voltage-gated calcium channels, and employed the transcript scanning method to identify alternatively spliced exons within the Ca V1.4 transcripts isolated from the human retina. In total, we identified 19 alternative splice variations, of which 16 variations have not been previously reported. Characterization of the C terminus alternatively spliced exons using whole-cell patch clamp electrophysiology revealed a splice variant that exhibits robust CDI. This splice variant arose from the splicing of a novel alternate exon (43*) that can be found in 13.6% of the full-length transcripts screened. Inclusion of exon 43*inserts a stop codon that truncates half the C terminus. The Ca V1.443*channel exhibited robust CDI, a larger current density, a hyperpolarized shift in activation potential by ∼10 mV, and a slower VDI. Through deletional experiments, we showed that the inhibitor of CDI was responsible for modulating channel activation and VDI, in addition to CDI. Calcium currents in the photoreceptors were observed to exhibit CDI and are more negatively activated as compared with currents elicited from heterologously expressed full-length Ca V1.4. Naturally occurring alternative splice variants may in part contribute to the properties of the native Ca V1.4 channels. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc.