Phenylarsine oxide (PAO)-mediated activation of phospholipase D in rat basophilic leukemia (RBL-2H3) cells: possible involvement of calcium and protein kinase C.
Addition of phenylarsine oxide (PAO) to [3H]oleic acid-labeled rat basophilic leukemia (RBL-2H3) cells gave rise to the remarkable formation of [3H]phosphatidylbutanol (PBut), a specific product of phospholipase D (PLD) activation. Preincubation of cells with 2,3-dimercaptopropanol (DMP) or dithiothreitol (DTT), compounds containing sulfhydryls, prevented PAO-stimulated [3H]PBut formation, indicating that PAO-stimulated PLD through interacting with vicinal thiol groups. Treatment of cells with PAO resulted in increase in intracellular Ca2+ concentration without significant production of inositol phosphates. Removal of extracellular free Ca2+ by chelating with EGTA was found to inhibit [3H]PBut formation by PAO. Incubation of cells with 20 nM phorbol 12-myristate 13-acetate (PMA) for 6 h caused down-regulation of protein kinase C (PKC) alpha and beta isozymes, whereas it had no effect on PKC delta, epsilon and zeta isozymes. Under this condition, decrease in PAO-stimulated [3H]PBut formation was observed to occur with a concomitant decrease in the level of PKC alpha and beta isozymes. These results suggest that a covalent bridge between vicinal thiol groups of cell surface proteins induced by PAO potentiates PLD activation and that PAO-induced PLD activation is regulated by Ca2+ and PKC alpha and/or beta isozymes.
Kumada
T
T
Department of Otolaryngology, Gifu University School of Medicine, Japan.
Nakashima
S
S
Nakamura
Y
Y
Miyata
H
H
Nozawa
Y
Y
D000818:Animals; D000941:Antigens / Q000276:immunology; D001152:Arsenicals / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D004112:Dimercaprol / Q000494:pharmacology; D004229:Dithiothreitol / Q000494:pharmacology; D004789:Enzyme Activation / Q000187:drug effects; D007946:Leukemia, Mast-Cell; D010739:Phospholipase D / Q000037:antagonists & inhibitors*; D011493:Protein Kinase C / Q000187:drug effects* / Q000378:metabolism*; D051381:Rats; D014407:Tumor Cells, Cultured
D000941:Antigens; D001152:Arsenicals; D004112:Dimercaprol; C029341:oxophenylarsine; D011493:Protein Kinase C; D010739:Phospholipase D; D002118:Calcium; D004229:Dithiothreitol