Phosphatidylinositol 3-kinase regulates glycosylphosphatidylinositol hydrolysis through PLC-gamma(2) activation in erythropoietin-stimulated cells.
Erythropoietin (Epo)-induced glycosylphosphatidylinositol (GPI) hydrolysis was previously described to be correlated with phospholipase C-gamma 2 (PLC-gamma2) activation. Here, we analyzed the involvement of phosphatidylinositol (PtdIns) 3-kinase in GPI hydrolysis through PLC-gamma2 tyrosine phosphorylation in response to Epo in FDC-P1 cells transfected with a wild type (WT) erythropoietin-receptor (Epo-R). We showed that phosphatidylinositol 3-kinase (PtdIns 3-kinase) inhibitor LY294002 inhibits Epo-induced hydrolysis of endogenous GPI and Epo-induced PLC-gamma2 tyrosine phosphorylation in a dose-dependent manner. Wortmannin, another PtdIns 3-kinase inhibitor, also suppressed Epo-induced PLC-gamma2 tyrosine phosphorylation. We also present evidence that PLC-gamma2 translocation to the membrane fraction on Epo stimulation is completely inhibited by LY294002. Upon Epo stimulation, the tyrosine-phosphorylated PLC-gamma2 was found to be associated with the tyrosine-phosphorylated Grb2-associated binder (GAB)2, SHC and SHP2 proteins. LY294002 cell preincubation did not affect GAB2, SHC and SHP2 tyrosine phosphorylation but inhibited the binding of PLC-gamma2 to GAB2 and SHP2. Taken together, these results show that PtdIns 3-kinase controls Epo-induced GPI hydrolysis through PLC-gamma2.
Laboratoire de Biochimie, CNRS, FRE 2534, IFR 53 Biomolécules, UFR Sciences Exactes et Naturelles, BP 1039, Université de Reims Champagne-Ardenne, 51687 Reims Cedex 2, France.
Cédric
C
Boudot
Zahra
Z
Kadri
Emmanuelle
E
Petitfrère
Elise
E
Lambert
Stany
S
Chrétien
Patrick
P
Mayeux
Bernard
B
Haye
Claudine
C
Billat
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D000818:Animals; D002454:Cell Differentiation / Q000187:drug effects / Q000502:physiology; D002455:Cell Division / Q000187:drug effects / Q000502:physiology; D002462:Cell Membrane / Q000187:drug effects / Q000201:enzymology*; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004912:Erythrocytes / Q000201:enzymology*; D015672:Erythroid Precursor Cells / Q000201:enzymology*; D004921:Erythropoietin / Q000235:genetics / Q000378:metabolism*; D017261:Glycosylphosphatidylinositols / Q000378:metabolism*; D015801:Helminth Proteins / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis / Q000187:drug effects; D007527:Isoenzymes / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D051966:Phospholipase C gamma; D010750:Phosphoproteins / Q000378:metabolism; D010766:Phosphorylation; D011485:Protein Binding / Q000502:physiology; D021381:Protein Transport / Q000187:drug effects / Q000502:physiology; D011506:Proteins / Q000378:metabolism; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D013347:Subcellular Fractions; D014162:Transfection; D010738:Type C Phospholipases / Q000378:metabolism*; D014443:Tyrosine / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; D033942:Adaptor Proteins, Vesicular Transport; D004791:Enzyme Inhibitors; C490895:GAB2 protein, human; D017261:Glycosylphosphatidylinositols; D015801:Helminth Proteins; D007527:Isoenzymes; D010750:Phosphoproteins; D011506:Proteins; C528024:SHC1 protein, human; D055765:Shc Signaling Adaptor Proteins; D000071425:Src Homology 2 Domain-Containing, Transforming Protein 1; D004921:Erythropoietin; C095048:microfilarial sheath protein, Helminth; D014443:Tyrosine; D019869:Phosphatidylinositol 3-Kinases; D010738:Type C Phospholipases; D051966:Phospholipase C gamma