The insulin-elicited 60-kDa phosphotyrosine protein in rat adipocytes is associated with phosphatidylinositol 3-kinase.
Insulin stimulates the tyrosine phosphorylation of a 60-kDa protein (pp60) in rat adipocytes. After insulin treatment of these cells, pp60, as well as the 160-kDa insulin receptor substrate-1 (IRS-1), were found to be associated with the enzyme phosphatidylinositol 3-kinase (PtdIns-3-kinase) in separate complexes. By contrast, pp60 was not detected in insulin-treated mouse 3T3-L1 adipocytes, which contain abundant IRS-1. PtdIns-3-kinase complex. The pp60.PtdIns 3-kinase complex was located in both the soluble and membrane fractions of the rat adipocytes. Fusion proteins containing the isolated src homology 2 domains from the 85-kDa subunit of PtdIns-3-kinase bound to pp60 in lysates of insulin-treated rat adipocytes. This finding indicates that the most likely mode of association of pp60 with PtdIns-3-kinase is through binding of phosphotyrosine residues in pp60 to these domains. By immunoaffinity chromatography on a monoclonal antibody against phosphotyrosine, pp60 was purified in high percentage yield from insulin-stimulated rat adipocytes, but the low amount of the protein obtained (about 3 ng from the adipocytes of one rat) precluded sequence analysis.
Lavan
B E
BE
Department of Biochemistry, Dartmouth Medical School, Hanover, New Hampshire 03755-3844.
Lienhard
G E
GE
D016475:3T3 Cells; D000273:Adipose Tissue / Q000378:metabolism*; D000818:Animals; D007328:Insulin / Q000378:metabolism*; D055504:Insulin Receptor Substrate Proteins; D051379:Mice; D019869:Phosphatidylinositol 3-Kinases; D010750:Phosphoproteins / Q000378:metabolism*; D010770:Phosphotransferases / Q000378:metabolism*; D019000:Phosphotyrosine; D051381:Rats; D017207:Rats, Sprague-Dawley; D014443:Tyrosine / Q000031:analogs & derivatives / Q000378:metabolism
D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526295:Irs1 protein, mouse; C526296:Irs1 protein, rat; D010750:Phosphoproteins; D019000:Phosphotyrosine; D014443:Tyrosine; D010770:Phosphotransferases; D019869:Phosphatidylinositol 3-Kinases