The epithelial sodium channel (ENaC) is intracellularly located as a tetramer.
The epithelial sodium channel (ENaC) plays an important role in Na(+) homeostasis by determining the Na(+) transport rate in so-called end-organs such as the renal collecting duct, distal colon, salivary and sweat gland ducts. ENaC is formed by heteromultimerization of three homologous subunits, termed alpha, beta, and gamma ENaC. The number of subunits and stoichiometry remain a matter of debate. In this study, sucrose gradient analysis of Xenopus laevis oocytes expressing rENaC revealed that ENaC forms heterotetramers, when the membrane fraction was solubilized in 0.1% (wt/vol) Na-deoxycholate. However, solubilization of the membrane proteins in higher concentrations of detergents dissociated the ENaC subunits of the tetramers in dimers. Co-immunoprecipitation studies with FLAG-tagged ENaC subunits suggest that during dissociation of ENaC tetramers the composition of dimers is completely random. Glycosidase digestion studies show that the ENaC subunits are retarded in the endoplasmic reticulum (ER) and pre-Golgi, whereas only a small fraction is inserted into the plasma membrane. Immunocytochemical analysis confirmed that ENaC is primarily located intracellularly. In addition, these findings are not restricted to the oocyte expression system, since identical results were found in rabbit connecting tubule and cortical collecting duct cells in primary culture and in rabbit colon.
Department of Cell Physiology, Nijmegen Centre for Molecular Life Sciences, University Medical Centre Nijmegen, The Netherlands.
Lisette
L
Dijkink
Anita
A
Hartog
Carel H
CH
van Os
René J M
RJ
Bindels
D000818:Animals; D002478:Cells, Cultured; D003106:Colon / Q000737:chemistry / Q000166:cytology; D004721:Endoplasmic Reticulum / Q000737:chemistry; D053503:Epithelial Sodium Channels; D006056:Golgi Apparatus / Q000737:chemistry; D015151:Immunoblotting; D007150:Immunohistochemistry; D007668:Kidney / Q000737:chemistry / Q000166:cytology; D009865:Oocytes / Q000737:chemistry / Q000166:cytology; D011233:Precipitin Tests; D020836:Protein Structure, Quaternary; D021122:Protein Subunits / Q000737:chemistry*; D011817:Rabbits; D015222:Sodium Channels / Q000737:chemistry*; D014982:Xenopus laevis
D053503:Epithelial Sodium Channels; D021122:Protein Subunits; D015222:Sodium Channels