Interactions between subunits of the human epithelial sodium channel

The human epithelial sodium channel (hENaC) mediates Na⁺ transport across the apical membrane of epithelia, and mutations in hENaC result in hypertensive and salt-wasting diseases. In heterologous expression systems, maximal hENaC function requires co-expression of three homologous proteins, the α, β, and γhENaC subunits, suggesting that hENaC subunits interact to form a multimeric channel complex. Using a co-immunoprecipitation assay, we found that hENaC subunits associated tightly to form homo- and heteromeric complexes and that the association between subunits occurred early in channel biosynthesis. Deletion analysis of γhENaC revealed that the N terminus was sufficient but not necessary for co-precipitation of αhENaC, and that both the N terminus and the second transmembrane segment (M2) were required for γ subunit function. The biochemical studies were supported by functional studies. Co-expression of γ subunits lacking M2 with full-length hENaC subunits revealed an inhibitory effect on hENaC channel function that appeared to be mediated by the cytoplasmic N terminus of γ, and was consistent with the assembly of nonfunctional subunits into the channel complex. We conclude that the N terminus of γhENaC is involved in channel assembly.