Insulin stimulates dephosphorylation of phosphorylase in rat epitrochlearis muscles.
We have investigated the effects of insulin on the phosphorylation of glycogen phosphorylase in skeletal muscle. Rat epitrochlearis muscles were incubated in vitro with 32Pi to label cellular phosphoproteins, before being treated with hormones. Phosphorylase, phosphorylase kinase, and glycogen synthase were immunoprecipitated under conditions that prevented changes in their phosphorylation states. Based on measurements of the activity ratio (-AMP/+AMP) and the 32P content of phosphorylase, 4-8% of the phosphorylase in untreated muscles appeared to be phosphorylated. Epinephrine promoted increases of approximately 4-fold in the 32P content and activity ratio. Neither these effects nor the epinephrine-stimulated increases in phosphorylation of glycogen synthase and phosphorylase kinase were attenuated by insulin. However, insulin at physiological concentrations rapidly decreased the 32P content of phosphorylase in muscles incubated without epinephrine. Results from peptide mapping experiments indicate that phosphorylase was phosphorylated at a single site in both control and insulin on phosphorylase represented a decrease in 32P of approximately 50%. By comparison, the 32P content of glycogen synthase and the beta subunit of phosphorylase kinase were decreased by only 20 and 16%, respectively; the 32P content of the kinase alpha subunit was not affected by insulin. The results provide direct evidence that insulin decreases the amount of phosphate in phosphorylase and phosphorylase kinase. These findings have important implications with respect to both the regulation of glycogen metabolism in skeletal muscle and the mechanism of insulin action.
Zhang
J N
JN
Department of Pharmacology, Washington University School of Medicine, St. Louis, Missouri 63110.
Hiken
J
J
Davis
A E
AE
Lawrence
J C
JC
D000818:Animals; D001665:Binding Sites; D004837:Epinephrine / Q000494:pharmacology; D006006:Glycogen Synthase / Q000378:metabolism; D007163:Immunosorbent Techniques; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D008297:Male; D008970:Molecular Weight; D009132:Muscles / Q000187:drug effects / Q000201:enzymology*; D010710:Phosphates / Q000378:metabolism; D010764:Phosphorylase Kinase / Q000378:metabolism; D006005:Phosphorylases / Q000378:metabolism*; D010766:Phosphorylation; D010768:Phosphoserine / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains
D007328:Insulin; D010710:Phosphates; D010768:Phosphoserine; D006005:Phosphorylases; D006006:Glycogen Synthase; D010764:Phosphorylase Kinase; D004837:Epinephrine