Dynamics of signaling during insulin-stimulated endocytosis of its receptor in adipocytes.
Insulin causes rapid insulin receptor autophosphorylation, receptor endocytosis, and phosphorylation of its principle substrate (IRS-1). Using rat adipocytes, we studied the dynamics of receptor autophosphorylation, the kinase activity, and the IRS-1 phosphorylation state relative to the subcellular localization of these proteins. After 2 min of insulin exposure, the specific phosphotyrosine content of the insulin receptor in the internal membranes (IM) peaks at a level 5-6-fold higher than the plasma membrane (PM) receptor and then declines after 5-8 min to a level similar to the PM receptor. The exogenous kinase activity of these receptors exactly mirrored their phosphotyrosine content. The distribution of IRS-1 is 80% cytosolic, 20% IM-associated, and essentially undetectable in the PM. The phosphorylation state of IRS-1 in the IM parallels that of the insulin receptor, but cytosolic IRS-1 phosphorylation remains constant. Insulin-dependent GLUT4 translocation to the PM occurs after the peak of IRS-1 phosphorylation. The data are consistent with the hypothesis that insulin action may be mediated by receptor internalization and interaction with its substrate(s) associated with internal membranes. A small fraction of phosphorylated insulin receptors is sufficient for signal transduction. The dephosphorylation of the insulin receptor and IRS-1 in the IM appears to be a concerted process, possibly mediated by the same enzyme.
Kublaoui
B
B
Department of Biochemistry, Boston University School of Medicine, Massachusetts 02118.
Lee
J
J
Pilch
P F
PF
D017667:Adipocytes / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology; D004705:Endocytosis / Q000187:drug effects*; D051275:Glucose Transporter Type 4; D007328:Insulin / Q000494:pharmacology*; D055504:Insulin Receptor Substrate Proteins; D007700:Kinetics; D008297:Male; D009004:Monosaccharide Transport Proteins / Q000378:metabolism; D009124:Muscle Proteins; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D051381:Rats; D017207:Rats, Sprague-Dawley; D011972:Receptor, Insulin / Q000187:drug effects / Q000378:metabolism*; D015398:Signal Transduction
D051275:Glucose Transporter Type 4; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; C526296:Irs1 protein, rat; D009004:Monosaccharide Transport Proteins; D009124:Muscle Proteins; D010750:Phosphoproteins; C495694:Slc2a4 protein, rat; D011972:Receptor, Insulin