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 alpha, beta, and gamma hENaC 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 home-and heteromeric complexes and that the association between subunits occurred early in channel biosynthesis. Deletion analysis of gamma hENaC revealed that the N terminus was sufficient but not necessary for co precipitation of alpha hENaC, and that both the N terminus and the second transmembrane segment (M2) were required for gamma subunit function. The biochemical studies were supported by functional studies. Go-expression of gamma 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 gamma, and was consistent with the assembly of nonfunctional subunits into the channel complex. We conclude that the N terminus of gamma hENaC is involved in channel assembly.