Biochemical composition of the functional amiloride‐sensitive, hetero‐multimeric, 4ps ENaC

Abstract

Epithelial sodium channels (ENaC) consist of α, β, and γ subunits. As ENaC subunit proteins are synthesized in the rough endoplasmic reticulum, they are core glycosylated and co‐assembled into a hero‐multimeric channel. As they traffic through the Golgi to the apical membrane, some subunits are further glycosylated in the Golgi because the sugars become resistant to endo‐H glycosidase digestion, a hallmark of Golgi glycosylation. Since the protease, furin is active in the trans‐Golgi and at the plasma membrane, but prostasins are active only at the plasma membrane, the ENaC α and γ subunits must be cleaved either in the trans‐Golgi or at the apical membrane or both. Apical membranes of renal A6 cells contain subunits from each maturation step, endo‐H‐resistant, endo‐H‐sensitive, cleaved, and un‐cleaved as determined by isolation of biotinylated apical subunits. Because A6 cells contain only one type of functional channel, functional channel must be formed only from a subset of these modified subunits corresponding to the different bands. Therefore, we analyzed which modified subunits make‐up functional channels using co‐immunoprecipitation and other techniques. This research is supported by DK‐037963.