Hydrogen peroxide-induced phospholipase D activation in rat pheochromocytoma PC12 cells: possible involvement of Ca2+-dependent protein tyrosine kinase.
The mechanism for hydrogen peroxide (H2O2)-induced phospholipase D (PLD) activation was investigated in [3H]palmitic acid-labeled PC12 cells. In the presence of butanol, H2O2 caused a great accumulation of [3H]phosphatidylbutanol in a concentration- or time-dependent manner. However, treatment with H2O2 of cell lysates exerted no effect on PLD activity. Treatment with H2O2 had only a marginal effect on phospholipase C (PLC) activation. A protein kinase C (PKC) inhibitor, Ro 31-8220, did not inhibit but rather slightly enhanced H2O2-induced PLD activity. Thus, H2O2-induced PLD activation is considered to be independent of the PLC-PKC pathway in PC12 cells. In contrast, pretreatment with tyrosine kinase inhibitor herbimycin A, genistein, or ST638 resulted in a concentration-dependent inhibition of H2O2-induced PLD activation. Western blot analysis revealed several apparent tyrosine-phosphorylated protein bands after the H2O2 treatment and tyrosine phosphorylation of these proteins was inhibited by these tyrosine kinase inhibitors. Moreover, depletion of extracellular Ca2+ abolished H2O2-induced PLD activation and protein tyrosine phosphorylation. Extracellular Ca2+ potentiated H2O2-induced PLD activation in a concentration-dependent manner. Taken together, these results suggest that a certain Ca2+-dependent protein tyrosine kinase(s) somehow participates in H2O2-induced PLD activation in PC12 cells.
Ito
Y
Y
Department of Biochemistry, Gifu University School of Medicine, Japan.
Nakashima
S
S
Nozawa
Y
Y
D000818:Animals; D016227:Benzoquinones; D015153:Blotting, Western; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002934:Cinnamates / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D019833:Genistein; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D007529:Isoflavones / Q000494:pharmacology; D047029:Lactams, Macrocyclic; D016716:PC12 Cells / Q000201:enzymology*; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D019000:Phosphotyrosine / Q000378:metabolism; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011809:Quinones / Q000494:pharmacology; D051381:Rats; D017828:Rifabutin / Q000031:analogs & derivatives; D013440:Sulfides / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010738:Type C Phospholipases / Q000378:metabolism
D016227:Benzoquinones; D002934:Cinnamates; D004791:Enzyme Inhibitors; D007529:Isoflavones; D047029:Lactams, Macrocyclic; D019269:Phosphatidylinositol 4,5-Diphosphate; D011809:Quinones; D013440:Sulfides; C060747:ST 638; D017828:Rifabutin; D019000:Phosphotyrosine; D000001:Calcimycin; C020754:herbimycin; D006861:Hydrogen Peroxide; D019833:Genistein; D011505:Protein-Tyrosine Kinases; D010738:Type C Phospholipases; D010739:Phospholipase D; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium