Effects of aldosterone on biosynthesis, traffic, and functional expression of epithelial sodium channels in A6 cells.
The collecting duct regulates Na(+) transport by adjusting the abundance/activity of epithelial Na(+) channels (ENaC). In this study we have investigated the synthesis, degradation, endocytosis, and activity of ENaC and the effects of aldosterone on these processes using endogenous channels expressed in the A6 cell line. Biochemical studies were performed with a newly raised set of specific antibodies against each of the three subunits of the amphibian ENaC. Our results indicate simultaneous transcription and translation of alpha, beta, and gamma subunits and enhancement of both processes by aldosterone: two- and fourfold increase, respectively. The biosynthesis of new channels can be followed by acquisition of endoglycosidase H-resistant oligosacharides in alpha and beta subunits and, in the case of alpha, by the appearance of a form resistant to reducing agents. The half-life of the total pool of subunits (t(1/2) 40-70 min) is longer than the fraction of channels in the apical membrane (t(1/2) 12-17 min). Aldosterone induces a fourfold increase in the abundance of the three subunits in the apical membrane without significant changes in the open probability, kinetics of single channels, or in the rate of degradation of ENaC subunits. Accordingly, the aldosterone response could be accounted by an increase in the abundance of apical channels due, at least in part, to de novo synthesis of subunits.
Alvarez de la Rosa
Diego
D
Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, CT 06510, USA.
Li
Hui
H
Canessa
Cecilia M
CM
D000450:Aldosterone / Q000494:pharmacology*; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism* / Q000502:physiology; D053503:Epithelial Sodium Channels; D006031:Glycosylation / Q000187:drug effects; D006207:Half-Life; D018408:Patch-Clamp Techniques; D021122:Protein Subunits; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D015222:Sodium Channels / Q000096:biosynthesis* / Q000235:genetics / Q000378:metabolism / Q000502:physiology*; D014158:Transcription, Genetic / Q000187:drug effects; D014982:Xenopus laevis
D053503:Epithelial Sodium Channels; D021122:Protein Subunits; D015222:Sodium Channels; D000450:Aldosterone