In vivo phosphorylation of the epithelial sodium channel.
The activity of the epithelial sodium channel (ENaC) in the distal nephron is regulated by an antidiuretic hormone, aldosterone, and insulin, but the molecular mechanisms that mediate these hormonal effects are mostly unknown. We have investigated whether aldosterone, insulin, or activation of protein kinases has an effect on the phosphorylation of the channel. Experiments were performed in an epithelial cell line generated by stable cotransfection of the three subunits (alpha, beta, and gamma) of ENaC. We found that beta and gamma, but not the alpha subunit, are phosphorylated in the basal state. Aldosterone, insulin, and protein kinases A and C increased phosphorylation of the beta and gamma subunits in their carboxyl termini, but none of these agents induced de novo phosphorylation of alpha subunits. Serines and threonines but not tyrosines were found to be phosphorylated. The results suggest that aldosterone, insulin, and protein kinases A and C modulate the activity of ENaC by phosphorylation of the carboxyl termini of the beta and gamma subunits.
Shimkets
R A
RA
Department of Genetics, Yale University School of Medicine, New Haven, CT 06520-8026, USA.
Lifton
R
R
Canessa
C M
CM
D000450:Aldosterone / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D005576:Colforsin / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D004285:Dogs; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D053503:Epithelial Sodium Channels; D007328:Insulin / Q000494:pharmacology; D008969:Molecular Sequence Data; D009399:Nephrons / Q000187:drug effects / Q000378:metabolism*; D010449:Peptide Mapping; D010748:Phosphopeptides / Q000302:isolation & purification; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D062687:Sodium Channel Agonists; D015222:Sodium Channels / Q000378:metabolism*; D014162:Transfection
D053503:Epithelial Sodium Channels; D007328:Insulin; D010748:Phosphopeptides; D062687:Sodium Channel Agonists; D015222:Sodium Channels; D005576:Colforsin; D000450:Aldosterone; D000584:Amiloride; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C