Open AccessInsulin-induced Stimulation of Na+,K+-ATPase Activity in Kidney Proximal Tubule Cells Depends on Phosphorylation of the α-Subunit at Tyr-10
Author(s) -
Eric Féraille,
Maria Luisa Carranza,
Sandrine Gonin,
Pascal Béguin,
Carlos H. Pedemonte,
M Rousselot,
Joseph Caverzasio,
Käthi Geering,
PierreYves Martin,
H Favre
Publication year - 1999
Publication title -
molecular biology of the cell
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.463
H-Index - 225
eISSN - 1939-4586
pISSN - 1059-1524
DOI - 10.1091/mbc.10.9.2847
Subject(s) - phosphorylation , tyrosine phosphorylation , biology , tyrosine , renal sodium reabsorption , protein tyrosine phosphatase , medicine , ouabain , reabsorption , endocrinology , convoluted tubule , insulin , biochemistry , kidney , sodium , chemistry , organic chemistry
Phosphorylation of the alpha-subunit of Na+,K(+)-ATPase plays an important role in the regulation of this pump. Recent studies suggest that insulin, known to increase solute and fluid reabsorption in mammalian proximal convoluted tubule (PCT), is stimulating Na+,K(+)-ATPase activity through the tyrosine phosphorylation process. This study was therefore undertaken to evaluate the role of tyrosine phosphorylation of the Na+,K(+)-ATPase alpha-subunit in the action of insulin. In rat PCT, insulin and orthovanadate (a tyrosine phosphatase inhibitor) increased tyrosine phosphorylation level of the alpha-subunit more than twofold. Their effects were not additive, suggesting a common mechanism of action. Insulin-induced tyrosine phosphorylation was prevented by genistein, a tyrosine kinase inhibitor. The site of tyrosine phosphorylation was identified on Tyr-10 by controlled trypsinolysis in rat PCTs and by site-directed mutagenesis in opossum kidney cells transfected with rat alpha-subunit. The functional relevance of Tyr-10 phosphorylation was assessed by 1) the abolition of insulin-induced stimulation of the ouabain-sensitive (86)Rb uptake in opossum kidney cells expressing mutant rat alpha1-subunits wherein tyrosine was replaced by alanine or glutamine; and 2) the similarity of the time course and dose dependency of the insulin-induced increase in ouabain-sensitive (86)Rb uptake and tyrosine phosphorylation. These findings indicate that phosphorylation of the Na+,K(+)-ATPase alpha-subunit at Tyr-10 likely participates in the physiological control of sodium reabsorption in PCT.