Mutation of Tyrosine 960 within the Insulin Receptor Juxtamembrane Domain Impairs Glucose Transport but Does Not Inhibit Ligand-mediated Phosphorylation of Insulin Receptor Substrate-2 in 3T3-L1 Adipocytes

• Published in: The Journal of biological chemistry 1999-04, Vol.274 (17), p.12075-12080
• Main Authors: Chaika, O V, Chaika, N, Volle, D J, Hayashi, H, Ebina, Y, Wang, L M, Pierce, J H, Lewis, R E
• Format: Article
• Language: English
• Subjects: 3T3 Cells; Adipocytes - metabolism; Animals; Biological Transport; Cell Membrane - metabolism; Cytoplasm - metabolism; Enzyme Activation; Glucose - metabolism; Insulin Receptor Substrate Proteins; Intracellular Signaling Peptides and Proteins; Ligands; Macrophage Colony-Stimulating Factor - pharmacology; Mice; Phosphatidylinositol 3-Kinases - metabolism; Phosphoproteins - metabolism; Phosphorylation; Receptor, Insulin - genetics; Receptor, Insulin - metabolism; Receptor, Macrophage Colony-Stimulating Factor - metabolism; Recombinant Fusion Proteins - metabolism; Tyrosine - genetics; Tyrosine - metabolism
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.274.17.12075

CSF-1 is equipotent to insulin in its ability to stimulate 2-[ 3 H]deoxyglucose uptake in 3T3-L1 adipocytes expressing the colony stimulating factor-1 receptor/insulin receptor chimera (CSF1R/IR). However, CSF-1-stimulated glucose uptake and glycogen synthesis is reduced by 50% in comparison to insulin in 3T3-L1 cells expressing a CSF1R/IR mutated at Tyr 960 (CSF1R/IRA960). CSF-1-treated adipocytes expressing the CSF1R/IRA960 were impaired in their ability to phosphorylate insulin receptor substrate 1 (IRS-1) but not in their ability to phosphorylate IRS-2. Immunoprecipitation of IRS proteins followed by Western blotting revealed that the intact CSF1R/IR co-precipitates with IRS-2 from CSF-1-treated cells. In contrast, the CSF1R/IRA960 co-precipitates poorly with IRS-2. These observations suggest that Tyr 960 is important for interaction of the insulin receptor cytoplasmic domain with IRS-2, but it is not essential to the ability of the insulin receptor tyrosine kinase to use IRS-2 as a substrate. These observations also suggest that in 3T3-L1 adipocytes, tyrosine phosphorylation of IRS-2 by the insulin receptor tyrosine kinase is not sufficient for maximal stimulation of receptor-regulated glucose transport or glycogen synthesis.