Post-translational palmitoylation controls the voltage gating and lipid raft association of CALHM1 channel

Abstract

Emerging roles of CALHM1, a recently discovered voltage-gated ion channel, include purinergic neurotransmission of tastes in taste buds and memory formation in the brain, highlighting its physiological importance. However, the regulatory mechanisms of the CALHM1 channel remain entirely unexplored, hindering full understanding of its contribution in vivo. In fact, the different gating properties of CALHM1 in vivo and in vitro suggest undiscovered regulatory mechanisms. Here, in searching for post-translational regulatory mechanisms, we discovered the regulation of CALHM1 gating and association with lipid microdomains via protein S-palmitoylation, the only reversible lipid modification of proteins on cysteine residues. CALHM1 is palmitoylated at two intracellular cysteines located in the juxtamembrane regions of the third and forth transmembrane domains. Enzymes that catalyze CALHM1 palmitoylation are identified by screening 23 members of the DHHC protein acyltransferase family. Epitope-tagging of endogenous CALHM1 proteins in mice reveals that CALHM1 is basally palmitoylated in taste buds in vivo. Functionally, palmitoylation downregulates CALHM1 without effects on its synthesis, degradation, and cell surface expression. Mutation of the palmitoylation sites has profound impact on CALHM1 gating, shifting the conductance-voltage relationship to more negative voltages and accelerating the activation kinetics. The same mutation also reduces CALHM1 association with detergent-resistant membranes. Our results comprehensively uncover a post-translational regulation of the voltage-dependent gating of CALHM1 by palmitoylation.

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