Insulin receptor function is inhibited by guanosine 5'-[gamma-thio]triphosphate (GTP[S]).
The regulatory role of GTP-binding proteins (G-proteins) in insulin receptor function was investigated using isolated insulin receptors and plasma membranes from rat adipocytes. Treatment of isolated insulin receptors with 1 mM-guanosine 5'-[gamma-thio]triphosphate (GTP[S]) inhibited insulin-stimulated phosphorylation of the beta-subunit, histone Hf2b and poly(GluNa4,Tyr1) by 22%, 65% and 65% respectively. Phosphorylation of calmodulin by the insulin receptor kinase was also inhibited by 1 mM-GTP[S] both in the absence (by 88%) and in the presence (by 81%) of insulin. In the absence of insulin, 1 mM-GTP had the same effect on calmodulin phosphorylation as 1 mM-GTP[S]. However, when insulin was present, GTP was less effective than GTP[S] (41% versus 81% inhibition). Concentrations of GTP[S] greater than 250 microM are necessary to inhibit phosphorylation. Although these concentrations are relatively high, the effect of GTP[S] is not due to competition with [32P]ATP for the insulin receptor kinase since (1) other nucleotide triphosphates did not inhibit phosphorylation as much as did GTP[S] (or GTP) and (2) the Vmax of the ATP-dependent kinase reaction was decreased in the presence of GTP[S]. GTP[S] (1 mM) also inhibited insulin binding to isolated receptors and plasma membranes, by 80% and 50% respectively. Finally, an antibody raised to a peptide sequence common to the alpha-subunits of G-proteins Gs, Gi, Go and transducin detected G-proteins in plasma membranes but failed to detect them in the insulin receptor preparation. These results indicate that GTP inhibits insulin receptor function, but does so through a mechanism that does not require a conventional GTP-binding protein.
Davis
H W
HW
Department of Pathology, Washington University School of Medicine, St. Louis, MO 63110.
McDonald
J M
JM
D000273:Adipose Tissue / Q000378:metabolism; D000818:Animals; D002147:Calmodulin / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism; D019204:GTP-Binding Proteins / Q000502:physiology*; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000494:pharmacology; D006657:Histones / Q000378:metabolism; D015151:Immunoblotting; D007328:Insulin / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins; D008297:Male; D010455:Peptides / Q000378:metabolism; D010766:Phosphorylation; D011505:Protein-Tyrosine Kinases / Q000378:metabolism; D051381:Rats; D011919:Rats, Inbred Strains; D011972:Receptor, Insulin / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013873:Thionucleotides / Q000494:pharmacology*
D002147:Calmodulin; D006657:Histones; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D010455:Peptides; D013873:Thionucleotides; C027606:L-glutamic acid-L-tyrosine copolymer; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006160:Guanosine Triphosphate; D011505:Protein-Tyrosine Kinases; D011972:Receptor, Insulin; D019204:GTP-Binding Proteins