scholarly journals Evidence for the calcium-dependent activation of phospholipase D in thrombin-stimulated human erythroleukaemia cells

1990 ◽  
Vol 267 (2) ◽  
pp. 479-483 ◽  
Author(s):  
S P Halenda ◽  
A G Rehm
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Phospholipase D ◽  
Calf Serum ◽  
Platelet Activating Factor ◽  
Diacylglycerol Kinase ◽  
Sole Source ◽  
Phospholipid Metabolism ◽  
Ionophore A23187 ◽  
Hel 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+.

Phosphatidic acid production in chitosan-elicited tomato cells, via both phospholipase D and phospholipase C/diacylglycerol kinase, requires nitric oxide

2011 ◽  
Vol 168 (6) ◽  
pp. 534-539 ◽  
Author(s):  
Nicolás Raho ◽  
Leonor Ramirez ◽  
M. Luciana Lanteri ◽  
Gabriela Gonorazky ◽  
Lorenzo Lamattina ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Phospholipase D ◽  
Acid Production ◽  
Diacylglycerol Kinase

scholarly journals Human platelet signaling defect characterized by impaired production of inositol-1,4,5-triphosphate and phosphatidic acid and diminished Pleckstrin phosphorylation: evidence for defective phospholipase C activation

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1676-1683 ◽  
Author(s):  
X Yang ◽  
L Sun ◽  
S Ghosh ◽  
AK Rao
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Platelet Activating Factor ◽  
Ionophore A23187 ◽  
Serotonin Secretion ◽  
Platelet Signaling ◽  
Intracellular Mediators ◽  
Gamma S ◽  
Hydrolysis Of ◽  
Pkc Activation

Signal transduction on platelet activation involves phosphoinositide- specific phospholipase C (PLC)-mediated hydrolysis of phosphatidylinositides and formation of inositol-1,4,5-triphosphate [I(1,4,5)P3], which mediates Ca2+ mobilization, and diacylglycerol (DG), which activates protein kinase C (PKC) to phosphorylate a 47-kD protein (Pleckstrin). We studied these events in two related patients previously reported (Blood 74:664, 1989) to have abnormal aggregation and 14C-serotonin secretion, and impaired intracellular Ca2+ mobilization in response to several agonists. Thrombin-induced I(1,4,5)P3 and phosphatidic acid formation were diminished. Pleckstrin phosphorylation was impaired on activation with thrombin, platelet- activating factor, and ionophore A23187, but was normal with PKC activator 1,2-dioctonyl-sn-glycerol (DiC8). Ca2+ mobilization induced by guanosine triphosphate (GTP) analog guanosine 5′-0-(3 thiotriphosphate) (GTP gamma S) was diminished. Pretreatment with either A23187 or DiC8 did not correct the impaired adenine diphosphate- induced secretion; however, upon stimulation with A23187 plus DiC8, pleckstrin phosphorylation and secretion were normal, indicating that both PKC activation and Ca2+ mobilization are essential for normal secretion. We conclude that these patients have a unique inherited platelet defect in formation of two key intracellular mediators [I(1,4,5)P3 and DG] and in the responses mediated by them due to a defect in postreceptor mechanisms of PLC activation.

scholarly journals Human platelet signaling defect characterized by impaired production of inositol-1,4,5-triphosphate and phosphatidic acid and diminished Pleckstrin phosphorylation: evidence for defective phospholipase C activation

Blood ◽  
1996 ◽  
Vol 88 (5) ◽  
pp. 1676-1683 ◽  
Author(s):  
X Yang ◽  
L Sun ◽  
S Ghosh ◽  
AK Rao
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Platelet Activating Factor ◽  
Ionophore A23187 ◽  
Serotonin Secretion ◽  
Platelet Signaling ◽  
Intracellular Mediators ◽  
Gamma S ◽  
Hydrolysis Of ◽  
Pkc Activation

Abstract Signal transduction on platelet activation involves phosphoinositide- specific phospholipase C (PLC)-mediated hydrolysis of phosphatidylinositides and formation of inositol-1,4,5-triphosphate [I(1,4,5)P3], which mediates Ca2+ mobilization, and diacylglycerol (DG), which activates protein kinase C (PKC) to phosphorylate a 47-kD protein (Pleckstrin). We studied these events in two related patients previously reported (Blood 74:664, 1989) to have abnormal aggregation and 14C-serotonin secretion, and impaired intracellular Ca2+ mobilization in response to several agonists. Thrombin-induced I(1,4,5)P3 and phosphatidic acid formation were diminished. Pleckstrin phosphorylation was impaired on activation with thrombin, platelet- activating factor, and ionophore A23187, but was normal with PKC activator 1,2-dioctonyl-sn-glycerol (DiC8). Ca2+ mobilization induced by guanosine triphosphate (GTP) analog guanosine 5′-0-(3 thiotriphosphate) (GTP gamma S) was diminished. Pretreatment with either A23187 or DiC8 did not correct the impaired adenine diphosphate- induced secretion; however, upon stimulation with A23187 plus DiC8, pleckstrin phosphorylation and secretion were normal, indicating that both PKC activation and Ca2+ mobilization are essential for normal secretion. We conclude that these patients have a unique inherited platelet defect in formation of two key intracellular mediators [I(1,4,5)P3 and DG] and in the responses mediated by them due to a defect in postreceptor mechanisms of PLC activation.

scholarly journals Stimulation of phosphatidate synthesis in endothelial cells in response to P2-receptor activation. Evidence for phospholipase C and phospholipase D involvement, phosphatidate and diacylglycerol interconversion and the role of protein kinase C

1992 ◽  
Vol 287 (1) ◽  
pp. 31-36 ◽  
Author(s):  
J R Purkiss ◽  
M R Boarder
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Phospholipase D ◽  
Purinergic Receptor ◽  
Diacylglycerol Kinase ◽  
Kinase C ◽  
Stimulation Of

To investigate the stimulation of phosphatidic acid formation in bovine aortic endothelial cells by P2-purinergic agonists, we labelled AG4762 cells with [32P]P1 and stimulated in the presence of butanol. Under these conditions phospholipase D generated [32P]phosphatidylbutanol, whereas the [32P]phosphatidic acid from phospholipase C and diacylglycerol kinase was unchanged. The action of various purinergic agonists on both [32P]phosphatidic acid and [32P]phosphatidylbutanol was consistent with the presence of a P2Y receptor. The stimulation of phospholipase D was dependent on extracellular Ca2+ and was mostly transient (completed within 3 min), whereas the initial stimulation of phospholipase C was independent of extracellular Ca2+, followed by a Ca(2+)-dependent phase. The agonist stimulation of phospholipase D was dependent on protein kinase C, as judged by its sensitivity to the relatively selective protein kinase C inhibitor Ro 31-8220. These results show that purinergic-receptor-mediated stimulation of phosphatidic acid has three phases: an initial Ca(2+)-independent stimulation of phospholipase C, an early but transient Ca(2+)- and protein kinase C-dependent stimulation of phospholipase D, and a sustained Ca(2+)-dependent stimulation of phospholipase C. Using propranolol to inhibit phosphatidate phosphohydrolase, we provide evidence that phosphatidic acid derived from purinergic-receptor-mediated stimulation of the phospholipase C/diacylglycerol kinase route can itself be converted back into diacylglycerol.

scholarly journals Comparison of diglyceride production from choline-containing phosphoglycerides in human neutrophils stimulated with N-formylmethionyl-leucylphenylalanine, ionophore A23187 or phorbol 12-myristate 13-acetate

1992 ◽  
Vol 286 (3) ◽  
pp. 693-699 ◽  
Author(s):  
M C Chabot ◽  
L C McPhail ◽  
R L Wykle ◽  
D A Kennerly ◽  
C E McCall
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Phospholipase D ◽  
Water Soluble ◽  
Incubation Mixture ◽  
Human Neutrophils ◽  
Ionophore A23187 ◽  
Agonist Stimulation

The turnover of choline-containing phosphoglycerides (PC) in response to agonist stimulation is well documented in human neutrophils. We have now compared the enzymic pathways of N-formylmethionyl-leucylphenylalanine (fMLP)-, A23187- and phorbol-12-myristate 13-acetate (PMA)-induced diglyceride (DG) and phosphatidic acid (PA) generation in these cells. In order to distinguish between phospholipase C- and D-mediated PC breakdown, human neutrophils were radiolabelled with 1-O-[3H]alkyl-2-acyl-glycero-3-phosphocholine and stimulated in the presence of ethanol or propranolol. The addition of 0.5% ethanol to the incubation mixture resulted in the production of phosphatidylethanol, indicative of phospholipase D activation, in response to all three stimuli. Concomitant with phosphatidylethanol formation was a partial block of PA production. The production of DG was also partially blocked by addition of ethanol. Propranolol (200 microM) was also used to assess the contributions of phospholipases C and D toward DG generation. Inhibition of PA phosphohydrolase by propranolol resulted in the complete abolition of DG generation when neutrophils were stimulated with fMLP. In contrast, propranolol only partially inhibited DG generation in response to A23187 and PMA. These results suggested that DG production in response to fMLP stimulation is mediated via the activation of phospholipase D, whereas A23187- or PMA-induced DG generation may involve more than one pathway. However, examination of the water-soluble choline metabolites produced indicated that phospholipase D was responsible for the production of PA and DG in response to all three stimuli.

scholarly journals The effects of phorbol ester, diacylglycerol, phospholipase C and Ca2+ ionophore on protein phosphorylation in human and sheep erythrocytes

1985 ◽  
Vol 232 (1) ◽  
pp. 43-47 ◽  
Author(s):  
P J Raval ◽  
D Allan
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cell Membrane ◽  
Phospholipase C ◽  
Phospholipid Metabolism ◽  
Ionophore A23187 ◽  
Myristate Acetate ◽  
Sheep Erythrocytes ◽  
Kinase C ◽  
Coomassie Blue

Treatment of human or sheep erythrocytes with PMA (phorbol myristate acetate) enhanced [32P]phosphate labelling of membrane polypeptides of approx. 100, 80 and 46 kDa. The 80 kDa and 46 kDa polypeptides coincided with bands 4.1 and 4.9 respectively on Coomassie-Blue-stained gels. Similar but smaller effects were obtained by treating human cells with 1-oleoyl-2-acetyl-rac-glycerol (OAG), exogenous bacterial phospholipase C or ionophore A23187 + Ca2+, each of which treatments would be expected to raise the concentration of membrane diacylglycerol. In contrast, sheep cells, which do not increase their content of diacylglycerol when treated with phospholipase C or A23187 + Ca2+, only showed enhanced phosphorylation with OAG. Neither human nor sheep cells showed any enhanced [32P]phosphate labelling of phosphoproteins when treated with 1-mono-oleoyl-rac-glycerol. It is concluded that diacylglycerol from a variety of sources can activate erythrocyte protein kinase C, but that the most effective diacylglycerol is that derived from endogenous polyphosphoinositides. In contrast with bacterial phospholipase C and A23187, which stimulate synthesis of phosphatidate by increasing the cell-membrane content of diacylglycerol in human erythrocytes, PMA, OAG or 1-mono-oleoyl-rac-glycerol caused no change in phospholipid metabolism.

Evidence for two mechanisms of thrombin-induced platelet activation: one proteolytic, one receptor mediated

1989 ◽  
Vol 67 (7) ◽  
pp. 332-336 ◽  
Author(s):  
Archibald McNicol ◽  
Jon M. Gerrard ◽  
D. Euan MacIntyre
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase C ◽  
Platelet Reactivity ◽  
Platelet Activating Factor ◽  
Receptor Activation ◽  
Acid Formation ◽  
Phosphoinositide Metabolism ◽  
Co Addition ◽  
The Individual ◽  
Dual Mechanism

The possibility that thrombin-induced platelet reactivity could occur via both a receptor-related and a proteolytic process was examined. Thrombin elicited the formation of considerably more [32P)phosphatidic acid (an index of phospholipase C catalysed phosphoinositide metabolism) than did platelet activating factor, 5-hydroxytryptamine, ADP, and the thromboxane A2 analogue EP171, when these agents were added either alone or in combination. Co-addition of thrombin and EP171 did not evoke significantly more [32P]phosphatidic acid than did thrombin alone. The protease inhibitor leupeptin, decreased but did not abolish [32P]phosphatidic acid formation elicited by either thrombin alone or thrombin in combination with EP171. The serine protease, trypsin, stimulated an increase in [32P]phosphatidic acid and this effect was additive with that of EP171. This augmentation by trypsin of EP171-induced [32P]phosphatidic acid formation was inhibited by leupeptin. These results are consistent with the concept that thrombin-induced activation of phospholipase C occurs by two distinct mechanisms: one via proteolysis, which is sensitive to leupeptin, and the other via receptor activation, a process shared by EP171. The individual components of this dual mechanism can be mimicked by the co-addition of a receptor-directed agonist (EP171) and a proteolytic agent (trypsin).Key words: platelet, thrombin, proteolysis, phosphoinositide.

scholarly journals Phospholipase D-derived phosphatidic acid is involved in the activation of the CD11b/CD18 integrin in human eosinophils

1999 ◽  
Vol 340 (1) ◽  
pp. 95-101 ◽  
Author(s):  
Anton T. J. TOOL ◽  
Michela BLOM ◽  
Dirk ROOS ◽  
Arthur J. VERHOEVEN
Keyword(s):  
Fatty Acid ◽  
Acid Phosphatase ◽  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Platelet Activating Factor ◽  
Induced Response ◽  
Acid Moiety ◽  
Kinase C ◽  
Phosphatidic Acid Phosphatase ◽  
A Minor

Priming of human eosinophils is an essential event for the respiratory burst induced by serum-opsonized particles [serum-treated zymosan (STZ)]. In this study we have found that treatment of eosinophils with platelet-activating factor (PAF) leads to activation of phospholipase D. Inhibition of the formation of phospholipase D-derived products by ethanol resulted in about 90% inhibition of PAF-induced binding of fluorescent STZ particles to the cells, but only when ethanol was added to the cells before treatment with PAF. When ethanol was added after treatment with PAF, only a minor inhibition of the STZ binding and STZ-induced response was observed. These results indicate that phospholipase D-derived phosphatidic acid is involved in PAF priming, without having an effect on STZ stimulation. In the presence of propranolol, which inhibits phosphatidic acid-phosphatase activity, binding of STZ particles to human eosinophils induced by suboptimal concentrations of PAF was enhanced, indicating that phosphatidic acid and not diradylglyceride is the relevant molecule derived from phospholipase D activity. Addition of cell-permeant diC8-phosphatidic acid (DiC8-PA) to human eosinophils resulted in CD11b/CD18-dependent adhesion, both to STZ particles and fibronectin-coated wells, without significant upregulation of CD11b/CD18. The DiC8-PA-induced adhesion was not mediated via the fatty acid moiety, because other C8-lipids such as 1,2-diC8-phosphatidylcholine, 1-C8-monoacylglycerol or C8-ceramide were without effect. Activation of protein kinase C with PMA or 1,2-diC8-diacylglycerol did result in enhanced STZ binding. However, under these latter conditions upregulation of CD11b/CD18 was observed. Taken together, these results suggest that phospholipase D-derived PA is involved in changing the affinity of the CD11b/CD18 integrin for its ligands.

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