Pertussis toxin inhibits phospholipase C activation and Ca2+ mobilization by sphingosylphosphorylcholine and galactosylsphingosine in HL60 leukemia cells. Implications of GTP-binding protein-coupled receptors for lysosphingolipids.
Extracellular sphingosylphosphorylcholine (SPC) and galactosylsphingosine (psychosine) induced Ca2+ mobilization in a dose-dependent manner in HL60 leukemia cells. The rapid and transient increase in intracellular Ca2+ concentration ([Ca2+]i) elicited by SPC and psychosine at concentrations lower than 30 microM was inhibited by treatment of the cells with pertussis toxin (PTX) and U73122, a phospholipase C inhibitor, as was the case for UTP, a P2-purinergic agonist. The increase in [Ca2+]i induced by these lysosphingolipids was associated with inositol phosphate production, which was also sensitive to PTX and U73122. The inositol phosphate response is not secondary to the increase in [Ca2+]i as evidenced by the observation that thapsigargin and ionomycin, Ca2+ mobilizing agents, never induced inositol phosphate production and, unlike lysosphingolipids, the [Ca2+]i rise by these agents was totally insensitive to PTX and U73122. When HL60 cells were differentiated into neutrophil-like cells by dibutyryl cyclic AMP, inositol phosphate and Ca2+ responses to AlF4- were enhanced, probably reflecting an increase in the amount of Gi2 and Gi3 compared with undifferentiated cells. In the neutrophil-like cells, however, the responses to SPC and psychosine were markedly attenuated. This may exclude the possibility that the lysosphingolipids activate rather directly PTX-sensitive GTP-binding proteins or the phospholipase C itself. Other lysosphingolipids including glucosylsphingosine (glucopsychosine) and sphingosylgalactosyl sulfate (lysosulfatides) at 30 microM or lower concentrations also showed PTX- and U73122-sensitive Ca2+ mobilization and inositol phosphate response in a way similar to SPC and psychosine. However, platelet-activating factor and lysoglycerophospholipids such as lysophosphatidylcholine and lysophosphatidic acid were less effective than these lysosphingolipids in the induction of Ca2+ mobilization. Taken together, the results indicate that a group of lysosphingolipids at appropriate doses induces Ca2+ mobilization through inositol phosphate production by phospholipase C activation. The lysosphingolipids-induced enzyme activation may be mediated by PTX-sensitive GTP-binding protein-coupled receptors, which may be different from previously identified platelet-activating factor receptor or lysophosphatidic acid receptor.
Okajima
F
F
Laboratory of Signal Transduction, Gunma University, Maebashi, Japan.
Kondo
Y
Y
D002118:Calcium / Q000378:metabolism*; D002454:Cell Differentiation; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D019204:GTP-Binding Proteins / Q000378:metabolism*; D018922:HL-60 Cells; D006801:Humans; D007700:Kinetics; D008246:Lysophospholipids / Q000494:pharmacology; D009504:Neutrophils / Q000166:cytology; D037342:Pertussis Toxin; D010767:Phosphorylcholine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010972:Platelet Activating Factor / Q000494:pharmacology; D011609:Psychosine / Q000494:pharmacology*; D011760:Pyrrolidinones / Q000494:pharmacology; D011956:Receptors, Cell Surface / Q000187:drug effects / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid; D013108:Sphingomyelin Phosphodiesterase / Q000494:pharmacology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014544:Uridine Triphosphate / Q000494:pharmacology; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*
D004791:Enzyme Inhibitors; D004963:Estrenes; D008246:Lysophospholipids; D010972:Platelet Activating Factor; D011760:Pyrrolidinones; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D049312:Receptors, Lysophospholipid; D010566:Virulence Factors, Bordetella; C005356:sphingosine phosphorylcholine; D010767:Phosphorylcholine; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; D011609:Psychosine; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D013108:Sphingomyelin Phosphodiesterase; D019204:GTP-Binding Proteins; D013110:Sphingosine; D002118:Calcium; D014544:Uridine Triphosphate