The voltage-gated sodium channel ß2 subunit associates with lipid rafts by S-palmitoylation.

ABSTRACT

The voltage-gated sodium channel is critical for cardiomyocyte function. It consists of a protein complex comprising a pore-forming α subunit and associated ß subunits. In polarized Madin-Darby canine kidney cells, we show evidence by acyl-biotin exchange that ß2 is S-acylated at Cys-182. Interestingly, we found that palmitoylation increases ß2 association with detergent-resistant membranes. ß2 localizes exclusively to the apical surface. However, depletion of plasma membrane cholesterol, or blocking intracellular cholesterol transport, caused mislocalization of ß2, as well as of the non-palmitoylable C182S mutant, to the basolateral domain. Apical ß2 did not undergo endocytosis and displayed limited diffusion within the plane of the membrane; such behavior suggests that, at least in part, it is cytoskeleton anchored. Upon acute cholesterol depletion, its mobility was greatly reduced, and a slight reduction was also measured as a result of lack of palmitoylation, supporting ß2 association with cholesterol-rich lipid rafts. Indeed, lipid raft labeling confirmed a partial overlap with apical ß2. Although ß2 palmitoylation was not required to promote surface localization of the α subunit, our data suggest that it is likely implicated in lipid raft association and the polarized localization of ß2.