Ca_Vβ controls the endocytic turnover of Ca_V1.2 L‐type calcium channel

Abstract

Membrane depolarization activates the multisubunit CaV1.2 L‐type calcium channel initiating various excitation coupling responses. Intracellular trafficking into and out of the plasma membrane regulates the channel's surface expression and stability, and thus, the strength of CaV1.2‐mediated Ca2+ signals. The mechanisms regulating the residency time of the channel at the cell membrane are unclear. Here, we coexpressed the channel core complex CaV1.2α1 pore‐forming and auxiliary CaVβ subunits and analyzed their trafficking dynamics from single‐particle‐tracking trajectories. Speed histograms obtained for each subunit were best fitted to a sum of diffusive and directed motion terms. The same mean speed for the highest‐mobility state underlying directed motion was found for all subunits. The frequency of this component increased by covalent linkage of CaVβ to CaV1.2α1 suggesting that high‐speed transport occurs in association with CaVβ. Selective tracking of CaV1.2α1 along the postendocytic pathway failed to show the highly mobile state, implying CaVβ‐independent retrograde transport. Retrograde speeds of CaV1.2α1 are compatible with myosin VI‐mediated backward transport. Moreover, residency time at the cell surface was significantly prolonged when CaV1.2α1 was covalently linked to CaVβ. Thus, CaVβ promotes fast transport speed along anterograde trafficking and acts as a molecular switch controlling the endocytic turnover of L‐type calcium channels.

ClC-3 regulates the excitability of nociceptive neurons and is involved in inflammatory processes within the spinal sensory pathway

“…Many ion channels are sorted via the secretory pathway ( Griffith, 2001 ; Swanwick et al, 2010 ; Capera et al, 2019 ) and recycled via clathrin-mediated endocytosis ( Okuse et al, 2002 ; Liu et al, 2005 ; Swanwick et al, 2010 ; Conrad et al, 2018 , 2021 ). ClC-3 has clathrin-binding motifs in its amino terminus ( Zhao et al, 2007 ; Stauber and Jentsch, 2010 ) and, thus, might facilitate the endocytosis of membrane proteins and direct them to lysosomes (ClC-3b) or the recycling endosome (ClC-3c) ( Guzman et al, 2015 , 2017 ; Comini et al, 2022 ).…”

Nanoscale architecture and dynamics of CaV1.3 channel clusters in cardiac myocytes revealed by single channel nanoscopy

“…Since we selectively labeled these tagged Ca V 1.3 variants and not endogenous Ca V 1.3 in our imaging experiments, simultaneous occurrence of both types of channels within clusters cannot be completely excluded. Offsetting this, we can assume that relative protein cell surface abundance was clearly weighted towards tagged Ca V 1.3 due to a titration effect: in contrast to endogenous channels, tagged Ca v 1.3 was overexpressed, while LTCC surface trafficking and residency depends on molecular assembly with β-subunits, limited by the available endogenous pool (Conrad et al, 2021; Stanika et al, 2016). In addition, ER-resident tagged Ca v 1.3 channels and aggregates were hardly observed in transfected cells, owing to a lower protein biosynthesis rate compared to heterologous expression systems and the sensitive unfolded protein response for ion channels in hiPSC-CM (Liu et al, 2018).…”

“…The pore-forming subunit α 1D was tagged at the N-terminus preserving channel voltage-gating, while the C-terminus is crucial for regulatory functions that may be perturbed by fusion-tagging. In addition, we considered that alternative tagging of the accessory β-subunit (Conrad et al, 2021; Del Villar et al, 2021; Liu et al, 2020) was unsuitable for our study since Ca V β can bind other Ca 2+ channel isoforms and performs intracellular functions (Vergnol et al, 2022). We developed independent and synergistic cluster analysis workflows on hiPSC-aCM expressing tagged Ca V 1.3 proteins.…”

The β2-Subunit of Voltage-Gated Calcium Channels Regulates Cardiomyocyte Hypertrophy

“…The physiological role of Ca v β has been assessed by several groups in different experimental models. In heterologous expression systems, Ca v β regulates LTCC membrane trafficking and Ca 2+ currents ( 6 , 7 ). In adult ventricular cardiomyocytes, a disruption of the Ca v 1.2-Ca v β association affects the inactivation rate of LTCC ( 8 ).…”