Sustained activation of phospholipase D via adenosine A3 receptors is associated with enhancement of antigen- and Ca(2+)-ionophore-induced secretion in a rat mast cell line.
The adenosine analog, N-ethylcarboxamidoadenosine (NECA), causes transient activation of phospholipase C and an enhancement of antigen-induced secretion in a rat mast cell (RBL-2H3) line via adenosine A3-receptors (Ramkumar et al., J. Biol. Chem. 268:16887, 1993) by a mechanism that is inhibited by bacterial toxins and potentiated by dexamethasone (Ali et al., J. Biol. Chem. 265:745-753, 1990). Here we show that NECA synergizes the secretory response to Ca(2+)-ionophore as well as to antigen. The ability of NECA to synergize the secretory responses persisted for 10 to 20 min, long after the early phospholipase C-mediated reactions to NECA had subsided. NECA caused, however, a dose-dependent sustained activation of phospholipase D, as indicated by the formation of [3H]phosphatidic acid, or in the presence of 0.3% ethanol, [3H]phosphatidylethanol. This activation was associated with a sustained increase in diglycerides, in protein kinase C activity and in the phosphorylation of myosin light chains by protein kinase C. The generation of diglycerides was enhanced in dexamethasone-treated cells and suppressed in cells that had been treated with cholera toxin or pertussis toxin. Collectively, the studies suggested that the generation of diglycerides via phospholipase D and the associated activation of protein kinase C were, by themselves, insufficient signals for secretion in RBL-2H3 cells, but that these reactions synergized responses to stimulants such as antigen or A23187 that caused substantial increases in [Ca2+]i.
Ali
H
H
Laboratory of Molecular Immunology, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
Choi
O H
OH
Fraundorfer
P F
PF
Yamada
K
K
Gonzaga
H M
HM
Beaven
M A
MA
D000241:Adenosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D019830:Adenosine-5'-(N-ethylcarboxamide); D000818:Animals; D000941:Antigens / Q000276:immunology; D000001:Calcimycin / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism; D002478:Cells, Cultured; D004075:Diglycerides / Q000096:biosynthesis; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation; D019204:GTP-Binding Proteins / Q000502:physiology; D007476:Ionophores / Q000494:pharmacology*; D008407:Mast Cells / Q000201:enzymology / Q000378:metabolism*; D010739:Phospholipase D / Q000378:metabolism*; D011493:Protein Kinase C / Q000378:metabolism; D058906:Purinergic P1 Receptor Agonists; D051381:Rats; D018047:Receptors, Purinergic P1 / Q000502:physiology*; D010738:Type C Phospholipases / Q000378:metabolism
D000941:Antigens; D004075:Diglycerides; D007476:Ionophores; D058906:Purinergic P1 Receptor Agonists; D018047:Receptors, Purinergic P1; D019830:Adenosine-5'-(N-ethylcarboxamide); D000001:Calcimycin; D011493:Protein Kinase C; D010738:Type C Phospholipases; D010739:Phospholipase D; D019204:GTP-Binding Proteins; D000241:Adenosine; D002118:Calcium