Role of p85 subunit of phosphatidylinositol-3-kinase as an adaptor molecule linking the insulin receptor, p62, and GTPase-activating protein.
After insulin stimulation of rat HTC hepatoma cells overexpressing normal human insulin receptors (IR), an antiserum to the p85 subunit of phosphatidylinositol-3-kinase (PIK) (alpha-p85) immunoprecipitated three major tyrosine-phosphorylated proteins: IR, insulin receptor substrate-1 (IRS-1), and a new 62-kDa protein (p62). Studies with antibodies to GTPase activating protein (alpha-GAP) and p62 ... GAP-associated protein suggested that p62 was the same as (or closely related to) p62 GAP-associated protein. In order to understand how p62 interacts with p85, we employed: 1) antibodies to the p110 subunit of PIK (alpha-p110); and 2) antiserum to IRS-1. To determine which subunit of PIK (p110 or p85) p62 associates with, we first immunoprecipitated insulin-treated cell lysates with alpha-p110 and subsequently immunoprecipitated with alpha-p85 followed by Western blotting analysis with anti-phosphotyrosine antibody (alpha-PY). In response to insulin, most of the tyrosine-phosphorylated p62 was complexed to p85 alone rather than with the PIK heterodimer. Moreover, p62 was absent in alpha-IRS-1 immunoprecipitates. These data suggest that: 1) p62 GAP-associated protein is tyrosine phosphorylated after insulin stimulation of cells; 2) p62 and IRS-1 form separate complexes with p85; 3) p62-GAP complex may be linked to p85 that is not bound to p110; 4) p85 may serve as an adaptor molecule in insulin receptor signaling, interacting with and regulating other intracellular proteins via SH2 domains.
Mesh Terms:
Animals, DNA-Binding Proteins, Enzyme Activation, GTPase-Activating Proteins, Humans, Immune Sera, Insulin Receptor Substrate Proteins, Kinetics, Peptide Fragments, Phosphatidylinositol 3-Kinases, Phosphoproteins, Phosphorylation, Phosphotransferases (Alcohol Group Acceptor), Precipitin Tests, Proteins, RNA-Binding Proteins, Rats, Receptor, Insulin, Signal Transduction, Tumor Cells, Cultured, Tyrosine
Animals, DNA-Binding Proteins, Enzyme Activation, GTPase-Activating Proteins, Humans, Immune Sera, Insulin Receptor Substrate Proteins, Kinetics, Peptide Fragments, Phosphatidylinositol 3-Kinases, Phosphoproteins, Phosphorylation, Phosphotransferases (Alcohol Group Acceptor), Precipitin Tests, Proteins, RNA-Binding Proteins, Rats, Receptor, Insulin, Signal Transduction, Tumor Cells, Cultured, Tyrosine
J. Biol. Chem.
Date: Apr. 29, 1994
PubMed ID: 8175658
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