CaBP1 and Calmodulin Compete in Regulating Calcium-Dependent Inactivation of CaV1.2

Abstract

CaV1.2 is an L-type calcium channel from a family of voltage dependent calcium channels (VDCC) distributed mainly in cardiac and smooth muscle, endocrine cells and neurons, which produce calcium influx in response to membrane depolarization. Interaction of calcium binding protein 1 (CaBP1) and calmodulin (CaM) with the C-terminus (CT) of the L-type CaV1.2 channel is crucial for calcium-dependent inactivation (CDI); CaBP1 has an opposite effect on VDCCs compared to CaM: while CaM promotes CDI, CaBP1 prolongs and facilitates calcium currents and does not support CDI. CaBP1 and CaM are also bind to the N-terminus (NT), but the role of this interactions and the effect of these proteins on each other are unknown. We characterized the interaction of CaBP1 with NT peptides of alpha-1C using pull down assay. This binding site of CaBP1 is between a.a. 95-120 and does not overlap the CaM binding site; the binding is not calcium dependent. Coexpression of CaBP1 with CaV1.2 in Xenopus oocytes eliminated the CaM-dependent CDI. Inversely, titrated expression of fluorescently tagged CaM and CaBP1 showed that CaM removed the effect of CaBP1 at a molar ratio of ∼a 1:1 in the cell. Alanine mutagenesis of the CaM binding site and deletion of the NT binding site of CaBP1 did not remove the effect of CaBP1 on the inactivation. Thus, NT does not appear to be the important anchoring site for the action CaBP1, and the role of the interaction of CaBP1 with the CaV1.2 NT remains to be determined.