A role for tyrosine phosphorylation in both activation and inhibition of the insulin receptor tyrosine kinase in vivo.
Upon insulin binding, a conformational change in the insulin receptor (IR) leads to IR beta-subunit autophosphorylation, an increase in IR beta-subunit exogenous tyrosine kinase activity, and the rapid endocytosis of the ligand-receptor complex into endosomes. Previous work has shown that upon internalization, rat hepatic endosomal IRs manifest increased autophosphorylating and exogenous tyrosine kinase activity compared to IRs located at the plasma membrane. As this period of enhanced activity is associated with reduced endosomal IR beta-subunit phosphotyrosine content, it has been proposed that partial dephosphorylation of the internalized IR beta-subunit by an endosomally located phosphotyrosine phosphatase(s) [PTPase(s)] mediates this effect. To test whether endosomal PTPase activity was required for internalization-dependent augmentation of IR tyrosine kinase activity, the present study used the peroxovanadium PTPase inhibitor, bisperoxo(1,10-phenanthroline)oxovanadate anion [bpV(phen)], to block IR dephosphorylation within this subcellular compartment. Rats were pretreated with bpV(phen) before receiving insulin (1.5 micrograms/100 g BW). bpV(phen) inhibited the dephosphorylation of 32P-labeled hepatic endosomal IR by approximately 97% at 15 min post-bpV(phen) injection and prevented a decrease in IR beta-subunit phosphotyrosine content after IR internalization. Fifteen-minute bpV(phen) pretreatment produced a significant reduction (75%; P < 0.001) in maximal insulin-stimulated endosomal IR exogenous kinase activity and decreased IR autophosphorylating activity by 4.3-fold in this subcellular fraction. In conclusion, these findings suggest that an hepatic endosomal PTPase(s) regulates internalization-dependent increases in IR exogenous tyrosine kinase activity.
Drake
P G
PG
Polypeptide Hormone Laboratory, McGill University, Montreal, Quebec, Canada.
Bevan
A P
AP
Burgess
J W
JW
Bergeron
J J
JJ
Posner
B I
BI
D000818:Animals; D002458:Cell Fractionation; D002462:Cell Membrane / Q000378:metabolism; D011992:Endosomes / Q000201:enzymology; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D007328:Insulin / Q000378:metabolism; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D046911:Macromolecular Substances; D009942:Organometallic Compounds / Q000494:pharmacology; D010618:Phenanthrolines / Q000494:pharmacology; D010766:Phosphorylation; D019000:Phosphotyrosine / Q000032:analysis; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011972:Receptor, Insulin / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
D004791:Enzyme Inhibitors; D007328:Insulin; D046911:Macromolecular Substances; D009942:Organometallic Compounds; D010618:Phenanthrolines; D019000:Phosphotyrosine; D014443:Tyrosine; C102120:bisperoxo(1,10-phenanthroline)oxovanadate(1-); D011972:Receptor, Insulin; D017027:Protein Tyrosine Phosphatases