Insulin treatment stimulates the tyrosine phosphorylation of the alpha-type 85-kDa subunit of phosphatidylinositol 3-kinase in vivo.
After adding insulin to cells overexpressing the insulin receptor, the activity of phosphatidylinositol (PI) 3-kinase in the anti-phosphotyrosine immunoprecipitates was rapidly and greatly increased. This enzyme may therefore be a substrate for the insulin receptor tyrosine kinase and may be one of the mediators of insulin signal transduction. However, it is unclear whether or not activated tyrosine kinase of the insulin receptor directly phosphorylates PI 3-kinase at tyrosine residue(s) and whether insulin stimulates the specific activity of PI 3-kinase. We reported previously that the 85-kDa subunit of purified PI 3-kinase was phosphorylated at tyrosine residue(s) by the insulin receptor in vitro. To examine the tyrosine phosphorylation of PI 3-kinase and change of its activity by insulin treatment in vivo, we used a specific antibody to the 85-kDa subunit of PI 3-kinase. The activity of PI 3-kinase in immunoprecipitates with the antibody against the p85 subunit of PI 3-kinase was increased about 3-fold by insulin treatment of cells overexpressing insulin receptors. Insulin treatment also stimulated the tyrosine, serine, and threonine phosphorylation of the alpha-type 85-kDa subunit of PI 3-kinase in vivo. Phosphatase treatment of the immunoprecipitates abolished the increase in PI 3-kinase activity. The phosphorylation(s) of the kinase itself, tyrosine phosphorylation(s) of associated protein(s), or the complex formation of the phosphorylated PI 3-kinase with associated proteins may increase the activity of PI 3-kinase.
Hayashi
H
H
Department of Enzyme Genetics, University of Tokushima, Japan.
Kamohara
S
S
Nishioka
Y
Y
Kanai
F
F
Miyake
N
N
Fukui
Y
Y
Shibasaki
F
F
Takenawa
T
T
Ebina
Y
Y
D000818:Animals; D016466:CHO Cells; D002522:Chlorocebus aethiops; D006224:Cricetinae; D007158:Immunologic Techniques; D066298:In Vitro Techniques; D007328:Insulin / Q000494:pharmacology*; D019869:Phosphatidylinositol 3-Kinases; D010766:Phosphorylation; D010770:Phosphotransferases / Q000276:immunology / Q000378:metabolism*; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin / Q000378:metabolism*; D011994:Recombinant Proteins / Q000378:metabolism; D013329:Structure-Activity Relationship; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D007328:Insulin; D019869:Phosphatidylinositol 3-Kinases; D010770:Phosphotransferases; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D011994:Recombinant Proteins; D014443:Tyrosine