Phosphatidylinositol kinase or an associated protein is a substrate for the insulin receptor tyrosine kinase.
The tyrosine kinase activity intrinsic to the insulin receptor is thought to be important in eliciting the intracellular responses to insulin; however, it has been difficult to determine the biochemical functions of the proteins which are substrates for this receptor. Treatment of Chinese hamster ovary (CHO) cells overexpressing the human insulin receptor (CHO.T) with insulin results in a 38 +/- 11 (mean +/- S.E., n = 9)-fold increase in a phosphatidylinositol (PtdIns) kinase activity in anti-phosphotyrosine immunoprecipitates of whole cell lysates. One minute of treatment of cells with insulin causes a dramatic increase in the PtdIns kinase activity in the anti-phosphotyrosine immunoprecipitates; the activity peaks within 5 min and remains elevated for at least 60 min after addition of insulin to the cells. This response is only slightly delayed compared with the time course we observe for activation of the insulin receptor tyrosine kinase. The insulin dose-response curves are also very similar for the activation of the insulin receptor tyrosine kinase activity and for the appearance of PtdIns kinase in the anti-phosphotyrosine immunoprecipitates. Stimulation of the endogenous insulin receptor of CHO cells also results in the association of PtdIns kinase activity with phosphotyrosine-containing proteins. However, CHO cells are less sensitive to insulin than CHO.T cells, and the maximal PtdIns kinase activity in antiphosphotyrosine immunoprecipitates from CHO cells is one-sixth that of CHO.T cells. In contrast, immunoprecipitates from CHO.T cells made with anti-insulin receptor antibodies do not contain significant levels of PtdIns kinase activity. This demonstrates that the PtdIns kinase is either a substrate for the insulin receptor tyrosine kinase or is tightly associated with another tyrosine phosphoprotein, which is not the insulin receptor.
Department of Cell Biology, Stanford University School of Medicine, California 94305-5332.
G
G
Endemann
K
K
Yonezawa
R A
RA
Roth
D019870:1-Phosphatidylinositol 4-Kinase; D000818:Animals; D002460:Cell Line; D006224:Cricetinae; D004305:Dose-Response Relationship, Drug; D006801:Humans; D007163:Immunosorbent Techniques; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D010770:Phosphotransferases / Q000378:metabolism*; D019000:Phosphotyrosine; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D011972:Receptor, Insulin; D014443:Tyrosine / Q000031:analogs & derivatives / Q000032:analysis / Q000378:metabolism
D007328:Insulin; D019000:Phosphotyrosine; D014443:Tyrosine; D010770:Phosphotransferases; D019870:1-Phosphatidylinositol 4-Kinase; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin