Insulin receptor tyrosine residues 1162 and 1163 control insulin stimulation of myristoyl-diacylglycerol generation and subsequent activation of glucose transport.
Chinese hamster ovary (CHO) transfectants expressing human insulin receptors that were mutated at tyrosines 1162 and 1163 (CHO-Y2 cells) exhibit decreased insulin stimulation of both receptor tyrosine kinase and 2-deoxyglucose uptake compared with transfectants expressing wild-type human insulin receptors (CHO-R cells). We now provide evidence that insulin stimulation of myristoyl-diacylglycerol (DAG) production is also markedly impaired in CHO-Y2 cells; this is manifested as a decreased responsiveness and sensitivity to insulin as compared with CHO-R and parental CHO cells. Further, we report that (i) the concentration-response curves of insulin-stimulated myristoyl-DAG production and 2-deoxyglucose uptake were superimposable within each of the three cell lines. (ii) The insulin-induced increase in myristoyl-DAG production preceded that in 2-deoxyglucose uptake, and the time course was altered for both responses in CHO-Y2 cells. (iii) Insulin also increased the phosphorylation of a 40-kDa protein known to be a substrate for protein kinase C, but to a much lesser extent in CHO-Y2 cells than in CHO-R cells. (iv) Exogenously added 1,2-dimyristoyl-glycerol and 4 beta-phorbol 12 beta-myristate 13 alpha-acetate (PMA) again stimulated both the phosphorylation of the 40-kDa protein and 2-deoxyglucose uptake, but in contrast to insulin, they elicited the same level of response in both CHO-R and CHO-Y2 cells. (v) Finally, in protein kinase C-depleted CHO-R cells, insulin and PMA stimulation of 40-kDa protein phosphorylation as well as PMA stimulation of 2-deoxyglucose uptake were completely abolished whereas insulin-stimulated 2-deoxyglucose uptake was only partially decreased. Taken together, these results suggest that insulin stimulation of 2-deoxyglucose uptake involves myristoyl-DAG production and, at least in part, protein kinase C activation, all three of these processes being controlled by receptor tyrosines 1162 and 1163.
Cherqui
G
G
Laboratoire de Biochimie, Institut National de la Santé et de la Rechetćhe Médicale Unité 181, Faculté de Médecine, Saint-Antoine, Paris, France.
Reynet
C
C
Caron
M
M
Melin
B
B
Wicek
D
D
Clauser
E
E
Capeau
J
J
Picard
J
J
D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D001693:Biological Transport, Active / Q000187:drug effects; D002460:Cell Line; D003847:Deoxyglucose / Q000378:metabolism*; D004075:Diglycerides / Q000096:biosynthesis* / Q000494:pharmacology; D007328:Insulin / Q000494:pharmacology*; D007700:Kinetics; D016297:Mutagenesis, Site-Directed; D010703:Phorbol Esters / Q000494:pharmacology; D010766:Phosphorylation; D011493:Protein Kinase C / Q000378:metabolism; D011972:Receptor, Insulin / Q000235:genetics* / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014162:Transfection; D014443:Tyrosine
D004075:Diglycerides; D007328:Insulin; D010703:Phorbol Esters; D014443:Tyrosine; C050054:dimyristoyl diglyceride; D000255:Adenosine Triphosphate; D003847:Deoxyglucose; D011972:Receptor, Insulin; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate