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Role of protein tyrosine phosphorylation in H2O2-induced activation of endothelial cell phospholipase D
V. Natarajan, S. Vepa, , W.M. Scribner
Published in American Physiological Society
PMID: 8843788
Volume: 271
Issue: 3 15-3
Pages: 400 - 408
Oxidant-induced activation of phospholipase D(PLD) in bovine pulmonary artery endothelial cells (BPAEC) is independent of protein kinase C and calcium. In the present study, the effects of tyrosine kinase and protein tyrosine phosphatase (PTPase) inhibitors on hydrogen peroxide (H2O2)- induced PLD activation and protein tyrosine phosphorylation were examined in BPAEC. Pretreatment of BPAEC with putative tyrosine kinase inhibitors genistein, tyrphostin, and herbimycin attenuated H2O2 (1 mM)-induced PLD activation. The inhibitory effect of the tyrosine kinase inhibitors was highly specific for H2O2-induced modulation and showed no effect on PLD activation mediated by 12-O-tetradecanoylphorbol 13-acetate or bradykinin. Furthermore, addition of H2O2 increased in a time-dependent manner tyrosine phosphorylation of several proteins (17 200 kDa), as determined by immunoblot analysis with antiphosphotyrosine antibodies. H2O2-mediated protein tyrosine phosphorylation preceded PLD activation, and a good correlation was observed on the effect of genistein in H2O2 induced PLD activation and protein tyrosine phosphorylation. Addition of vanadate, a phosphotyrosine phosphatase inhibitor, synergistically increased both PLD activation and protein tyrosine phosphorylation mediated by H2O2. Moreover, vanadate by itself had minimal effect on basal PLD activity in BPAEC; however, at 10 μM vanadate, an increase in protein tyrosine phosphorylation was observed. In addition to vanadate, phenylarsine oxide and diamide potentiated H2O2- induced PLD activation. These results suggest that tyrosine kinase activation may be involved in H2O2-induced PLD activation in vascular endothelial cells.
About the journal
JournalAmerican Journal of Physiology - Lung Cellular and Molecular Physiology
PublisherAmerican Physiological Society