Effect of GTP gamma S on insulin binding and tyrosine phosphorylation in liver membranes and L6 muscle cells.
Guanosine 5'-O-(3-thiotriphosphate) (GTP gamma S), a specific activator of G proteins, did not change the Kd nor total binding of [125I]insulin in plasma membranes from rat liver. Insulin did not alter GTP gamma 35S binding nor polypeptide ADP ribosylation in crude and plasma membranes catalyzed either intrinsically or by cholera toxin. In L6 muscle cells, insulin caused tyrosine phosphorylation of a polypeptide of Mr 160,000. Cell electroporation enabled testing of G protein action in this cellular system. Phosphorylation of the Mr 160,000 polypeptide in these permeabilized cells was insulin and ATP dependent but other small molecules or ionic gradients were not essential. The reaction could not be mimicked by the G protein agonist GTP gamma S nor inhibited by the G protein antagonist guanosine 5'-O-(2-thiodiphosphate) (GDP beta S). However, GTP gamma S effectively decreased insulin-mediated phosphorylation of this polypeptide. This suggests that the tyrosine kinase activity of the insulin receptor can be modulated by G protein agonists. It is concluded that cross talk between the insulin receptor and G proteins could not be demonstrated in isolated membranes by strategies that detect interactions between beta-adrenergic receptors and G proteins. In contrast, in permeabilized cells, G protein-mediated regulation of the insulin receptor kinase activity could be detected.
Burdett
E
E
Department of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
Mills
G B
GB
Klip
A
A
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism; D002463:Cell Membrane Permeability; D002772:Cholera Toxin / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D006153:Guanosine Diphosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D006160:Guanosine Triphosphate / Q000031:analogs & derivatives* / Q000378:metabolism / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008297:Male; D009132:Muscles / Q000378:metabolism; D009243:NAD / Q000378:metabolism; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D011972:Receptor, Insulin / Q000187:drug effects / Q000378:metabolism*; D013873:Thionucleotides / Q000378:metabolism / Q000494:pharmacology*; D014443:Tyrosine
D007328:Insulin; D013873:Thionucleotides; D009243:NAD; D006153:Guanosine Diphosphate; D000246:Adenosine Diphosphate Ribose; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D014443:Tyrosine; C023427:guanosine 5'-O-(2-thiodiphosphate); D006160:Guanosine Triphosphate; D002772:Cholera Toxin; D011972:Receptor, Insulin; D019204:GTP-Binding Proteins