Evidence for the calcium-dependent activation of phospholipase D in thrombin-stimulated human erythroleukaemia cells.
Human erythroleukaemia (HEL) cells were exposed to thrombin and other platelet-activating stimuli, and changes in radiolabelled phospholipid metabolism were measured. Thrombin caused a transient fall in PtdInsP and PtdInsP2 levels, accompanied by a rise in diacylglycerol and phosphatidic acid, indicative of a classical phospholipase C/diacylglycerol kinase pathway. However, the rise in phosphatidic acid preceded that of diacylglycerol, which is inconsistent with phospholipase C/diacylglycerol kinase being the sole source of phosphatidic acid. In the presence of ethanol, thrombin and other agonists (platelet-activating factor, adrenaline and ADP, as well as fetal-calf serum) stimulated the appearance of phosphatidylethanol, an indicator of phospholipase D activity. The Ca2+ ionophore A23187 and the protein kinase C activator phorbol myristate acetate (PMA) also elicited phosphatidylethanol formation, although A23187 was at least 5-fold more effective than PMA. Phosphatidylethanol production stimulated by agonists or A23187 was Ca2(+)-dependent, whereas that with PMA was not. These result suggest that phosphatidic acid is generated in agonist-stimulated HEL cells by two routes: phospholipase C/diacylglycerol kinase and phospholipase D. Activation of the HEL-cell phospholipase D in response to agonists may be mediated by a rise in intracellular Ca2+.
Halenda
S P
SP
Department of Pharmacology, University of Missouri School of Medicine, Columbia 65212.
Rehm
A G
AG
D000244:Adenosine Diphosphate / Q000494:pharmacology; D000001:Calcimycin / Q000494:pharmacology; D002118:Calcium / Q000494:pharmacology*; D002460:Cell Line; D004789:Enzyme Activation; D004837:Epinephrine / Q000494:pharmacology; D020404:Glycerophospholipids; D006801:Humans; D007700:Kinetics; D004915:Leukemia, Erythroblastic, Acute; D019814:Myristic Acid; D009227:Myristic Acids / Q000378:metabolism; D010712:Phosphatidic Acids / Q000378:metabolism; D010716:Phosphatidylinositols / Q000378:metabolism; D010739:Phospholipase D / Q000378:metabolism*; D010740:Phospholipases / Q000378:metabolism*; D010972:Platelet Activating Factor / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology / Q000031:analogs & derivatives; D013917:Thrombin / Q000494:pharmacology*; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000201:enzymology*
D020404:Glycerophospholipids; D009227:Myristic Acids; D010712:Phosphatidic Acids; D010716:Phosphatidylinositols; D010972:Platelet Activating Factor; C051521:phosphatidylethanol; D019814:Myristic Acid; D000001:Calcimycin; C016658:phorbolol myristate acetate; D000244:Adenosine Diphosphate; D010740:Phospholipases; D010739:Phospholipase D; D013917:Thrombin; D013755:Tetradecanoylphorbol Acetate; D002118:Calcium; D004837:Epinephrine