Platelet-derived growth factor-induced H(2)O(2) production requires the activation of phosphatidylinositol 3-kinase.
Autophosphorylation of the platelet-derived growth factor (PDGF) receptor triggers intracellular signaling cascades as a result of recruitment of Src homology 2 domain-containing enzymes, including phosphatidylinositol 3-kinase (PI3K), the GTPase-activating protein of Ras (GAP), the protein-tyrosine phosphatase SHP-2, and phospholipase C-gamma1 (PLC-gamma1), to specific phosphotyrosine residues. The roles of these various effectors in PDGF-induced generation of H(2)O(2) have now been investigated in HepG2 cells expressing various PDGF receptor mutants. These mutants included a kinase-deficient receptor and receptors in which various combinations of the tyrosine residues required for the binding of PI3K (Tyr(740) and Tyr(751)), GAP (Tyr(771)), SHP-2 (Tyr(1009)), or PLC-gamma1 (Tyr(1021)) were mutated to Phe. PDGF failed to increase H(2)O(2) production in cells expressing either the kinase-deficient mutant or a receptor in which the two Tyr residues required for the binding of PI3K were replaced by Phe. In contrast, PDGF-induced H(2)O(2) production in cells expressing a receptor in which the binding sites for GAP, SHP-2, and PLC-gamma1 were all mutated was slightly greater than that in cells expressing the wild-type receptor. Only the PI3K binding site was alone sufficient for PDGF-induced H(2)O(2) production. The effect of PDGF on H(2)O(2) generation was blocked by the PI3K inhibitors LY294002 and wortmannin or by overexpression of a dominant negative mutant of Rac1. These results suggest that a product of PI3K is required for PDGF-induced production of H(2)O(2) in nonphagocytic cells, and that Rac1 mediates signaling between the PI3K product and the putative NADPH oxidase.
Bae
Y S
YS
Center for Cell Signaling Research, Division of Molecular Life Sciences, and Department of Biological Sciences, Ewha Womans University, Seoul 120-750, Korea. baeys@mm.ewha.ac.kr
Sung
J Y
JY
Kim
O S
OS
Kim
Y J
YJ
Hur
K C
KC
Kazlauskas
A
A
Rhee
S G
SG
D001665:Binding Sites; D006528:Carcinoma, Hepatocellular; D004789:Enzyme Activation; D006801:Humans; D006861:Hydrogen Peroxide / Q000378:metabolism*; D008113:Liver Neoplasms; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010766:Phosphorylation; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology*; D020797:Receptor, Platelet-Derived Growth Factor beta / Q000737:chemistry / Q000187:drug effects / Q000502:physiology*; D011994:Recombinant Proteins / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D014162:Transfection; D014407:Tumor Cells, Cultured
D010982:Platelet-Derived Growth Factor; D011994:Recombinant Proteins; D006861:Hydrogen Peroxide; D019869:Phosphatidylinositol 3-Kinases; D020797:Receptor, Platelet-Derived Growth Factor beta