Designer genetically encoded voltage‐dependent calcium channel inhibitors inspired by RGK GTPases

Abstract

High-voltage-activated calcium (Ca V 1/Ca V 2) channels translate action potentials into Ca 2+ influx in excitable cells to control essential biological processes that include; muscle contraction, synaptic transmission, hormone secretion and activity-dependent regulation of gene expression. Modulation of Ca V 1/Ca V 2 channel activity is a powerful mechanism to regulate physiology, and there are a host of intracellular signalling molecules that tune different aspects of Ca V channel trafficking and gating for this purpose. Beyond normal physiological regulation, the diverse Ca V channel modulatory mechanisms may potentially be co-opted or interfered with for therapeutic benefits. Ca V 1/Ca V 2 channels are potently inhibited by a four-member sub-family of Ras-like GTPases known as RGK (Rad, Rem, Rem2, Gem/Kir) proteins. Understanding the mechanisms by which RGK proteins inhibit Ca V 1/Ca V 2 channels has led to the development of novel genetically encoded Ca V channel blockers with unique properties; including, chemoand optogenetic control of channel activity, and blocking channels either on the basis of their Henry M. Colecraft, PhD, is the John C. Dalton Professor of Physiology and Cellular Biophysics at Columbia University Vagelos College of Physicians and Surgeons. Dr Colecraft's laboratory focuses on modulation of voltage-gated ion channels (by intracellular signalling proteins, auxiliary subunits and posttranslational modifications) and understanding molecular/biophysical mechanisms underlying diseases caused by ion channel mutations (ion channelopathies). His lab has used protein engineering approaches to develop genetically encoded molecules to inhibit or modulate the activity of ion channels for customized applications.This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited. 1684 H. M. Colecraft J Physiol 598.9subcellular localization or by targeting an auxiliary subunit. These genetically encoded Ca V channel inhibitors have outstanding utility as enabling research tools and potential therapeutics.Abstract figure legend RGK proteins are small Ras-like GTPases that potently inhibit voltage-gated calcium (CaV) channels by binding their auxiliary b subunits. Mechanistic insights into how RGK proteins inhibit CaV channels has been exploited to develop novel genetically-encoded CaV channel inhibitors that can be acutely activated by small molecules or light, or produce constitutive inhibition via targeted ubiquitination using CaVb-binding nanobodies. Advantages of such genetically-encoded CaV channel inhibitors include their ability to be selectively targeted to specific tissue, cell types, sub-cellular localization, and distinct CaV channel macromolecular complexes.

Role of High Voltage-Gated Ca2+ Channel Subunits in Pancreatic β-Cell Insulin Release. From Structure to Function

“…ELKS binding to β 2 subunit in pancreatic β-cells enhanced L-type Ca 2+ current at the vascular side of the β-cell plasma membrane and regulated the initial polarized Ca 2+ influx and rapid insulin vesicle exocytosis [152]. Another important mechanism that dynamically modulates the HVCC membrane incorporation and function is the interaction of the β and α 1 subunits with the small GTPases RGK protein family members (Rem, Rem2, Rad, Gem/Kir) [153]. Initially it has been shown that co-expression of Gem with Ca V 1.2 or Ca V 1.3 channels resulted in Ca 2+ influx inhibition, and these findings were later extended to the other RGK family members (Rad [154], Rem and Rem2 [155,156], Gem [157,158]) and HVCC isoforms [156,159].…”

Inhibitory effects on L- and N-type calcium channels by a novel CaVβ1 variant identified in a patient with autism spectrum disorder

“…Of note, this was also seen in differences between Gem-mediated inhibitory effects. RGK-proteins have been proposed to have potentials as therapeutic tools for a differential and specific modulation of VGCCs (Colecraft 2020 ). It is tempting to speculate that RGK proteins might be a tool to differentially compensate for changes of VGCC-mediated currents that are due to mutated Ca V β subunits.…”

The life cycle of voltage-gated Ca2+ channels in neurons: an update on the trafficking of neuronal calcium channels

“…The development of genetically encoded Ca V channel inhibitors has been one of the foci of the Colecraft lab for many years [ 145 ]. Understanding the mechanisms by which RGK proteins inhibit Ca V channels has allowed them to engineer RGK proteins that specifically inhibit subtypes of Ca V channels in cardiomyocytes and in neurons, i.e.…”