Intermolecular phosphorylation of insulin receptor as possible mechanism for amplification of binding signal

• Published in: Diabetes (New York, N.Y.) Vol. 40; no. 2; pp. 300 - 303
• Main Authors: HAYES, G. R, LYDON, L. D, LOCKWOOD, D. H
• Format: Journal Article
• Language: English
• Published: Alexandria, VA American Diabetes Association 01.02.1991
• Subjects: Animals; Antibodies, Monoclonal - immunology; Biological and medical sciences; Cell Membrane - metabolism; Cell Membrane - physiology; Cell Membrane - ultrastructure; Cell receptors; Cell structures and functions; Female; Fundamental and applied biological sciences. Psychology; Hormone receptors. Growth factor receptors. Cytokine receptors. Prostaglandin receptors; Humans; Insulin - metabolism; Insulin - pharmacology; Liver - cytology; Liver - physiology; Liver - ultrastructure; Molecular and cellular biology; Phosphorylation; Placenta - cytology; Placenta - physiology; Placenta - ultrastructure; Precipitin Tests; Pregnancy; Protein Binding - physiology; Rats; Receptor, Insulin - immunology; Receptor, Insulin - metabolism; Receptor, Insulin - physiology; Signal Transduction - physiology; Signal transduction; Pancreatic hormone; Phosphorylation; Binding site; Insulin; Hormonal receptor
• ISSN: 0012-1797; 1939-327X
• DOI: 10.2337/diab.40.2.300

Clustering of cell-surface insulin receptors has led to the speculation that intermolecular phosphorylation of unoccupied receptors catalyzed by ligand-occupied receptors within the cluster could be a mechanism by which the insulin-binding signal is amplified. We examined whether insulin receptors can be phosphorylated by an intermolecular mechanism. In this study, we used highly purified insulin receptors isolated from rat liver plasma membranes and human placental membranes. Rat liver insulin receptors were "activated" by incubation with 10 nM insulin in the presence of ATP. Subsequent to removal of insulin by immunodepletion, these receptors were used as an enzyme source to study phosphorylation of unphosphorylated "substrate" human receptors. Initially, we found no evidence that the addition of activated rat receptors increased phosphorylation of human receptors, when assessed by immunoprecipitation with a human-specific monoclonal antibody. To examine the possibility that these negative results were due to insufficient receptor concentration, activated human receptors were mixed with unphosphorylated substrate receptors at concentrations up to 60 micrograms/ml. In this study, we found that addition of activated receptors resulted in increased phosphorylation of the substrate receptors at the highest concentrations employed. These are the first data indicating that insulin receptors per se are capable of intermolecular phosphorylation. In vivo, this could be the initial step in amplifying the insulin-binding signal.