Insulin- and glucose-induced phosphorylation of the Na(+),K(+)-adenosine triphosphatase alpha-subunits in rat skeletal muscle.
Phosphorylation of the alpha-subunits of Na(+),K(+)-adenosine triphosphatase in response to insulin, high extracellular glucose concentration, and phorbol 12-myristate 13-acetate was investigated in isolated rat soleus muscle. All three stimuli increased alpha-subunit phosphorylation approximately 3-fold. Phorbol 12-myristate 13-acetate- and high glucose-induced phosphorylation of the alpha-subunit was completely abolished by the PKC inhibitor GF109203X, whereas insulin-stimulated phosphorylation was only partially reduced. Notably, insulin stimulation resulted in phosphorylation of the alpha-subunit on serine, threonine, and tyrosine residues, whereas high extracellular glucose or phorbol 12-myristate 13-acetate stimulation mediated phosphorylation only on serine and threonine residues. Insulin stimulation resulted in translocation of Na(+),K(+)-adenosine triphosphatase alpha(2)-subunit to the plasma membrane and increased Na(+),K(+)-adenosine triphosphatase activity in the same membrane fraction. High glucose had no effect on alpha-subunits distribution. Immunoprecipitation with antiphosphotyrosine antibody and subsequent Western blot analysis with anti-alpha(1)- and -alpha(2)-subunit antibodies revealed that both alpha(1)- and alpha(2)-subunit isoforms underwent phosphorylation on tyrosine residues in response to insulin, although with different time course and magnitude. Thus, we show that insulin-stimulated phosphorylation of Na(+),K(+)-adenosine triphosphatase alpha-subunit occurs via a PKC- and tyrosine kinase-dependent mechanism, whereas high glucose-induced phosphorylation is only PKC-dependent. Phosphorylation of Na(+),K(+)-adenosine triphosphatase alpha-subunits may be involved in regulation of Na(+),K(+)-adenosine triphosphatase activity by insulin or high extracellular glucose in skeletal muscle.
Chibalin
A V
AV
Department of Clinical Physiology, Karolinska Hospital, 171 76 Stockholm, Sweden. alexander.chibalin@kirurgi.ki.se
Kovalenko
M V
MV
Ryder
J W
JW
Féraille
E
E
Wallberg-Henriksson
H
H
Zierath
J R
JR
D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005110:Extracellular Space / Q000378:metabolism; D005947:Glucose / Q000378:metabolism / Q000494:pharmacology*; D007211:Indoles / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D008297:Male; D008301:Maleimides / Q000494:pharmacology; D018482:Muscle, Skeletal / Q000187:drug effects / Q000201:enzymology*; D019606:Phosphoamino Acids / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D017208:Rats, Wistar; D000254:Sodium-Potassium-Exchanging ATPase / Q000378:metabolism*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014018:Tissue Distribution / Q000187:drug effects; D014443:Tyrosine / Q000378:metabolism
D004791:Enzyme Inhibitors; D007211:Indoles; D007328:Insulin; D008301:Maleimides; D019606:Phosphoamino Acids; D014443:Tyrosine; D000254:Sodium-Potassium-Exchanging ATPase; D005947:Glucose; C070515:bisindolylmaleimide I; D013755:Tetradecanoylphorbol Acetate