MOLECULAR DIVERSITY AND FUNCTIONAL-CHARACTERIZATION OF VOLTAGE-DEPENDENT CALCIUM CHANNELS (CACN4) EXPRESSED IN PANCREATIC BETA-CELLS

Dihydropyridine-sensitive voltage-dependent calcium channels (VDCC) pl ay a crucial role in insulin secretion. We recently have cloned a huma n alpha(1)-subunit of the VDCC expressed in pancreatic beta-cells, des ignated CACN4. In this study we have isolated complementary DNAs encod ing two forms of rat CACN4 (rCACN4A and rCACN4B) from a rat insulinoma RINm5F complementary DNA library. Rat CACN4A is a protein of 2203 ami no acids and is the rat homolog of human CACN4, whereas rCACN4B lacks 535 amino acids in the carboxyl-terminal region, probably due to alter native splicing. We have found two additional variations, one in the i ntracellular loop between repeats I and II and the other in the extrac ellular region between the third and fourth segments of repeat IV. Rev erse transcriptase-polymerase chain reaction analysis of rat pancreati c islet messenger RNA reveals that these variants are present in pancr eatic islets. In addition, whole-cell voltage-damp recordings of Chine se hamster ovary cells stably expressing the alpha(1)-subunit (rCACN4A or rCACN4B) with or without the calcium channel beta(2)-subunit show that coexpression of rCACN4A with the beta(2)-subunit or rCACN4B with the beta(2)-subunit elicits L-type VDCC currents, whereas expression o f the alpha(1)-subunit alone does not, indicating that CACN4 can assoc iate functionally with the beta(2)-subunit and that the beta-subunit i s essential for functional expression of CACN4. These results suggest that there are various subtypes of CACN4 expressed in pancreatic beta- cells, and that both rCACN4A and rCACN4B can function as VDCC. Further more, the present study suggests that the expression of the beta-subun it as well as the alpha(1)-subunit may participate in the regulation o f insulin secretion.