Mapping the interaction surface between CaVβ and actin and its role in calcium channel clearance

Defective ion channel turnover and clearance of damaged proteins are associated with aging and neurodegeneration. The L-type Ca_V1.2 voltage-gated calcium channel mediates depolarization-induced calcium signals in heart and brain. Here, we determined the interaction surface between actin and two calcium channel subunits, Ca_Vβ₂ and Ca_Vβ₄, using cross-linking mass spectrometry and protein-protein docking, and uncovered a role in replenishing conduction-defective Ca_V1.2 channels. Computational and in vitro mutagenesis identified hotspots in Ca_Vβ that decreased the affinity for actin but not for Ca_V1.2. When coexpressed with Ca_V1.2, none of the tested actin-association-deficient Ca_Vβ mutants altered the single-channel properties or the total number of channels at the cell surface. However, coexpression with the Ca_Vβ₂ hotspot mutant downregulated current amplitudes, and with a concomitant reduction in the number of functionally available channels, indicating that current inhibition resulted from a build-up of conduction silent channels. Our findings established Ca_Vβ₂-actin interaction as a key player for clearing the plasma membrane of corrupted Ca_V1.2 proteins to ensure the maintenance of a functional pool of channels and proper calcium signal transduction. The Ca_Vβ-actin molecular model introduces a potentially druggable protein-protein interface to intervene Ca_V-mediated signaling processes.