The molecular basis of the inhibition of CaV1 calcium-dependent inactivation by the distal carboxy tail

Abstract

Ca 2+ /calmodulin-dependent inactivation (CDI) of Ca V channels is a critical regulatory process that tunes the kinetics of Ca 2+ entry for different cell types and physiologic responses. CDI is mediated by calmodulin (CaM), which is bound to the IQ domain of the Ca V carboxy tail. This modulatory process is tailored by alternative splicing such that select splice variants of Ca V 1.3 and Ca V 1.4 contain a long distal carboxy tail (DCT). The DCT harbors an inhibitor of CDI (ICDI) module that competitively displaces CaM from the IQ domain, thereby diminishing CDI. While this overall mechanism is now well described, the detailed interactions required for ICDI binding to the IQ domain are yet to be elucidated. Here, we perform alanine-scanning mutagenesis of the IQ and ICDI domains and evaluate the contribution of neighboring regions to CDI inhibition. Through FRET binding analysis, we identify functionally relevant residues within the Ca V 1.3 IQ domain and the Ca V 1.4 ICDI and nearby A region, which are required for high-affinity IQ/ICDI binding. Importantly, patch-clamp recordings demonstrate that disruption of this interaction commensurately diminishes ICDI function resulting in the re-emergence of CDI in mutant channels. Furthermore, Ca V 1.2 channels harbor a homologous DCT; however, the ICDI region of this channel does not appear to appreciably modulate Ca V 1.2 CDI. Yet coexpression of Ca V 1.2 ICDI with select Ca V 1.3 splice variants significantly disrupts CDI, implicating a cross-channel modulatory scheme in cells expressing both channel subtypes. In all, these findings provide new insights into a molecular rheostat that fine-tunes Ca 2+ -entry and supports normal neuronal and cardiac function.

Calmodulin promotes a Ca2+‐dependent conformational change in the C‐terminal regulatory domain of CaV1.2

“…This lack of CaM binding to Ca V 1.4 is caused by a downstream domain in Ca V 1.4 (called ICDI) [26] which disables CaM binding. However, removal of the ICDI domain allows Ca V 1.4 to interact with CaM and restore CDI [27,28]. Consistent with these previous results, we find in our current study that CT(1455–1604) CaV1.4 (that lacks the ICDI domain) is structurally similar to CT(1520–1669) CaV1.2 (Fig.…”

Alzheimer’s Disease beyond Calcium Dysregulation: The Complex Interplay between Calmodulin, Calmodulin-Binding Proteins and Amyloid Beta from Disease Onset through Progression

“…CDI is a widely used ion-channel regulatory mechanism that involves both apoCaM and Ca 2+ /CaM binding to cytoplasmic domains close to the channel pore opening [ 36 , 37 ]. During normal calcium homeostasis, apoCaM binds to an IQ-like domain in the C-terminal of NMDAR, enhancing channel opening [ 94 ]. When intracellular calcium levels rise, the resident apoCaM is transformed to Ca 2+ /CaM, leading to its repositioning at the Ca 2+ -dependent CaMBD.…”

Regulation of Cardiac Cav1.2 Channels by Calmodulin

“…CDI of Ca 2+ channels was subsequently reported in various eukaryotic cell types across kingdoms, including placozoans, poriferans, ciliates, and vertebrates [ 112 , 113 ]. Among Cav channels in vertebrates, Cav1.2, Cav1.3, and Cav2.1 (P/Q type) channels show definitive CDI, while Cav1.1 shows modest CDI, Cav2.2 (N type) and Cav2.3 (R type) channels show tiny or undetectable CDI, and Cav1.4 and Cav3 (low-voltage activated type) channels show vestigial CDI [ 114 , 115 ].…”