AJP - Cell Physiology, Vol 258, Issue 1 C99-108, Copyright © 1990 by American Physiological Society
Effect of GTP gamma S on insulin binding and tyrosine phosphorylation in liver membranes and L6 muscle cells
E. Burdett, G. B. Mills and A. Klip
Department of Cell Biology, Hospital for Sick Children, Toronto, Ontario, Canada.
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.
