Mechanism of tyrosine phosphorylation and activation of phospholipase C-gamma 1. Tyrosine 783 phosphorylation is not sufficient for lipase activation.
Phospholipase C-gamma 1 (PLC-gamma 1) is phosphorylated on three tyrosine residues: Tyr-771, Tyr-783, and Tyr-1253. With the use of antibodies specific for each of these phosphorylation sites, we have now determined the kinetics and magnitude of phosphorylation at each site. Phosphorylation of Tyr-783, which is essential for lipase activation, was observed in all stimulated cell types examined. The extent of phosphorylation of Tyr-1253 was approximately 50 to 70% of that of Tyr-783 in cells stimulated with platelet-derived growth factor (PDGF) or epidermal growth factor (EGF), but Tyr-1253 phosphorylation was not detected in B or T cell lines stimulated through B- and T-cell antigen receptors, respectively. Tyr-771 was phosphorylated only at a low level in all cells studied. In cells stimulated with PDGF, phosphorylation and dephosphorylation of Tyr-783 and of Tyr-1253 occurred with similar kinetics; the receptor kinase appeared to phosphorylate both sites, albeit with Tyr-783 favored over Tyr-1253, before the bound PLC-gamma 1 was released, and phosphorylation at the two sites occurred independently. PDGF and EGF induced similar levels of phosphorylation of Tyr-783 and of Tyr-1253 in a cell line that expressed receptors for both growth factors. However, only PDGF, not EGF, elicited substantial PLC activity, suggesting that Tyr-783 phosphorylation was not sufficient for enzyme activation. Finally, concurrent production of phosphatidylinositol 3,4,5-trisphosphate was found to contribute to the activation of phosphorylated PLC-gamma 1.
Sekiya
Fujio
F
Laboratory of Cell Signaling, National Heart, Lung, and Blood Institute, National Institutes of Health, Bethesda, Maryland 20892, USA.
Poulin
Benoit
B
Kim
Yeun Ju
YJ
Rhee
Sue Goo
SG
D000818:Animals; D000906:Antibodies / Q000737:chemistry; D001402:B-Lymphocytes / Q000378:metabolism; D001665:Binding Sites; D002455:Cell Division; D002460:Cell Line; D004789:Enzyme Activation; D004815:Epidermal Growth Factor / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D015151:Immunoblotting; D007295:Inositol Phosphates / Q000378:metabolism; D019169:Jurkat Cells; D007700:Kinetics; D008049:Lipase / Q000378:metabolism*; D051379:Mice; D008957:Models, Genetic; D009154:Mutation; D041681:NIH 3T3 Cells; D051966:Phospholipase C gamma; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011233:Precipitin Tests; D051381:Rats; D013601:T-Lymphocytes / Q000378:metabolism; D013997:Time Factors; D010738:Type C Phospholipases / Q000737:chemistry* / Q000378:metabolism; D014443:Tyrosine / Q000737:chemistry / Q000378:metabolism*
D000906:Antibodies; D007295:Inositol Phosphates; D010982:Platelet-Derived Growth Factor; D014443:Tyrosine; D004815:Epidermal Growth Factor; D008049:Lipase; D010738:Type C Phospholipases; D051966:Phospholipase C gamma