scholarly journals The temporal relationship between phospholipase activation, diradylglycerol formation and superoxide production in the human neutrophil

1990 ◽  
Vol 271 (1) ◽  
pp. 209-213 ◽  
Author(s):  
N T Thompson ◽  
J E Tateson ◽  
R W Randall ◽  
G D Spacey ◽  
R W Bonser ◽  
...  
Keyword(s):  
Phospholipase D ◽  
Human Neutrophil ◽  
Second Messengers ◽  
Superoxide Production ◽  
Human Neutrophils ◽  
Inositol 1,4,5 Trisphosphate ◽  
Phosphatidate Phosphohydrolase ◽  
Time Courses ◽  
Rapid Activation ◽  
Stimulation Of

Fluctuations in the amounts of choline, inositol 1,4,5-trisphosphate (IP3) and diradylglycerol have been used to monitor phospholipase activation in the human neutrophil. Stimulation of human neutrophils by formylmethionyl-leucylphenylalanine (fMet-Leu-Phe) resulted in a rapid activation of both phosphatidylinositol 4,5-bisphosphate breakdown by phospholipase C and phosphatidylcholine breakdown by phospholipase D. Diradylglycerol accumulation occurred more slowly than that of either choline or IP3 and was inhibited by 30 mM-butanol, suggesting that the bulk was derived from the phospholipase D pathway via phosphatidate phosphohydrolase. Consistent with this is the observation that choline and diradylglycerol are produced in similar amounts. 1,2-Diacylglycerol (DAG) and 1-O-alkyl-2-acyl-sn-glycerol species accumulated with different time courses, indicating that one or more steps in the phospholipase D pathway was selective for the diacyl species. Superoxide production by fMet-Leu-Phe-stimulated neutrophils paralleled DAG accumulation over the first 5 min, but thereafter this production stopped, despite the fact that DAG remained elevated. We conclude that DAG derived from the phospholipase D pathway is only one of the second messengers important in controlling this functional response.

scholarly journals ADP-ribosylation-factor-regulated phospholipase D activity localizes to secretory vesicles and mobilizes to the plasma membrane following N-formylmethionyl-leucyl-phenylalanine stimulation of human neutrophils

1997 ◽  
Vol 325 (3) ◽  
pp. 581-585 ◽  
Author(s):  
C. P. MORGAN ◽  
H. SENGELOV ◽  
J. WHATMORE ◽  
N. BORREGAARD ◽  
S. COCKCROFT
Keyword(s):  
Plasma Membrane ◽  
Phospholipase D ◽  
Small Gtpases ◽  
Human Neutrophils ◽  
Secretory Vesicles ◽  
Rho Proteins ◽  
Adp Ribosylation ◽  
Activated Neutrophils ◽  
Adp Ribosylation Factor ◽  
Stimulation Of

Phospholipase D (PLD) is responsible for the hydrolysis of phosphatidylcholine to produce phosphatidic acid and choline. Human neutrophils contain PLD activity which is regulated by the small GTPases, ADP-ribosylation factor (ARF) and Rho proteins. In this study we have examined the subcellular localization of the ARF-regulated PLD activity in non-activated neutrophils and cells ‘primed‘ with N-formylmethionyl-leucyl-phenylalanine (fMetLeuPhe). We report that PLD activity is localized at the secretory vesicles in control cells and is mobilized to the plasma membrane upon stimulation with fMetLeuPhe. We conclude that the ARF-regulated PLD activity is translocated to the plasma membrane by secretory vesicles upon stimulation of neutrophils with fMetLeuPhe in inflammatory/priming doses. We propose that this relocalization of PLD is important for the subsequent events occurring during neutrophil activation.

Regulation of p21rac Activation in Human Neutrophils

Blood ◽  
1999 ◽  
Vol 94 (3) ◽  
pp. 1121-1130 ◽  
Author(s):  
Niels Geijsen ◽  
Sanne van Delft ◽  
Jan A.M. Raaijmakers ◽  
Jan-Willem J. Lammers ◽  
John G. Collard ◽  
...  
Keyword(s):  
G Protein ◽  
Kinase Inhibitors ◽  
Platelet Activating Factor ◽  
Superoxide Production ◽  
Mitogen Activated Protein Kinase ◽  
Human Neutrophils ◽  
Gm Csf ◽  
Mitogen Activated Protein ◽  
Macrophage Colony ◽  
Stimulation Of

The small guanosine triphosphate (GTPase) p21rac is highly expressed in human neutrophils where it is thought to play a role in cytoskeletal reorganization and superoxide production. Using the p21rac binding domain of PAK (PAK-RBD) as an activation-specific probe, we have investigated agonist-stimulated activation of p21rac. Stimulation of neutrophils with the chemoattractants fMet-Leu-Phe (fMLP) or platelet-activating factor (PAF) induced an extremely rapid and transient p21rac activation, being optimal within 5 seconds. This activation correlates with the rapid changes of intracellular free Ca2+ ([Ca2+]i) stimulated by fMLP; however, changes in [Ca2+]i were neither sufficient nor required for p21rac activation. Furthermore, fMLP-induced p21rac activation was not inhibited by broad tyrosine kinase inhibitors or specific inhibitors of ERK, p38 mitogen activated protein kinase, Src, or phosphatidylinositol 3-kinases. Surprisingly, the cytokines granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumor necrosis factor- did not cause p21rac activation or modulate fMLP-induced p21rac activation. AlF−, a potent activator of heterotrimeric G-protein -subunits, however, was found to activate p21rac. Stimulation of neutrophils with phorbol myristate acetate (PMA) strongly activated the respiratory burst, but did not induce p21rac activation, suggesting that superoxide production per se can occur independently of p21rac activation. These data suggest that in human granulocytes, G-protein coupled receptors, but not cytokine receptors, activate p21rac via a rapid, novel exchange-mechanism independently of changes in [Ca2+]i, tyrosine phosphorylation, or PI3K.

scholarly journals Tyrosine phosphorylation is involved in receptor coupling to phospholipase D but not phospholipase C in the human neutrophil

1992 ◽  
Vol 281 (3) ◽  
pp. 597-600 ◽  
Author(s):  
I J Uings ◽  
N T Thompson ◽  
R W Randall ◽  
G D Spacey ◽  
R W Bonser ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Tyrosine Kinase ◽  
Phospholipase C ◽  
Tyrosine Phosphorylation ◽  
Phospholipase D ◽  
Human Neutrophil ◽  
Human Neutrophils ◽  
Receptor Coupling ◽  
Kinase C

The tyrosine kinase inhibitors ST271, ST638 and erbstatin inhibited phospholipase D (PLD) activity in human neutrophils stimulated by fMet-Leu-Phe, platelet-activating factor and leukotriene B4. These compounds did not inhibit phorbol ester-stimulated PLD, indicating that they do not inhibit PLD per se, but probably act at a site between the receptor and the phospholipase. In contrast, the protein kinase C inhibitor Ro-31-8220 inhibited phorbol 12,13-dibutyrate- but not fMet-Leu-Phe-stimulated PLD activity, arguing against the involvement of protein kinase C in the receptor-mediated activation of PLD. ST271 did not inhibit Ins(1,4,5)P3 generation, but did inhibit protein tyrosine phosphorylation stimulated by fMet-Leu-Phe. The phosphotyrosine phosphatase inhibitor pervanadate increased tyrosine phosphorylation and stimulated PLD. These results suggest that tyrosine kinase activity is involved in receptor coupling to PLD but not to PtdIns(4,5)P2-specific phospholipase C in the human neutrophil.

scholarly journals Phospholipase D activation is functionally linked to superoxide generation in the human neutrophil

1989 ◽  
Vol 264 (2) ◽  
pp. 617-620 ◽  
Author(s):  
R W Bonser ◽  
N T Thompson ◽  
R W Randall ◽  
L G Garland
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Human Neutrophil ◽  
Superoxide Production ◽  
Superoxide Generation ◽  
Inositol 1 ◽  
Opsonized Zymosan

Neutrophils stimulated with formylmethionyl-leucylphenylalanine (fMet-Leu-Phe) in the presence of butanol and ethanol formed phosphatidyl alcohols through a phospholipase D mechanism. The alcohols inhibited phosphatidic acid and diradylglycerol (DRG) formation, but did not block inositol 1, 4, 5-trisphosphate release. fMet-Leu-Phe-stimulated superoxide production was inhibited by alcohol concentrations which blocked DRG formation, whereas opsonized-zymosan-stimulated superoxide production was only partially decreased. These results suggest that phospholipase D activation is functionally linked to superoxide production in the human neutrophil.

scholarly journals Staurosporine clamps cytosolic free Ca2+ concentrations of human neutrophils

1992 ◽  
Vol 283 (2) ◽  
pp. 499-505 ◽  
Author(s):  
K Wong ◽  
L Kwan-Yeung ◽  
J Turkson
Keyword(s):  
Binding Assay ◽  
Radioligand Binding ◽  
Human Neutrophils ◽  
Radioligand Binding Assay ◽  
Permeabilized Cells ◽  
Inositol 1,4,5 Trisphosphate ◽  
Combination Studies ◽  
Peptide Stimulation ◽  
Linear Manner ◽  
Stimulation Of

The present studies indicate that 50 nM-10 microM-staurosporine increased cytosolic free Ca2+ concentrations ([Ca2+]i) of fura-2-loaded neutrophils in a non-linear manner. The rise in [Ca2+]i was rapid, reaching a plateau (e.g. to 0.4 microM with 1 microM-staurosporine) within 30 s, and was maintained for more than 20 min. Pretreating cells with pertussis toxin had no effect on this reaction. The elevation of [Ca2+]i was insensitive to extracellular Ca2+ concentrations and was due entirely to mobilization of intracellular Ca2+ stores. Mn(2+)-quench studies confirmed the absence of Ca2+ influx. No Ca2+ efflux occurred in staurosporine-treated cells. In combination studies, staurosporine potentiated Ca2+ influx induced by N-formylmethionyl-leucyl-phenylalanine (FMLP) and did not block Ca2+ efflux associated with peptide stimulation of neutrophils. Studies with permeabilized cells showed that staurosporine did not directly release intracellular Ca2+ stores, nor did it affect the sequestration of Ca2+ by a Ca2+/ATPase pump. A radioligand-binding assay failed to detect changes in the level of inositol 1,4,5-trisphosphate in neutrophils incubated with less than or equal to 1 microM-staurosporine, but in cells treated with 10 microM-staurosporine the assay recorded a transient increase in this second messenger similar to that induced by FMLP. Finally, lysozyme, but not beta-glucuronidase, was released from staurosporine-treated cells. The present results suggest that staurosporine increased [Ca2+]i by indirectly mobilizing internal Ca2+ stores. Staurosporine suppression of Ca2+ efflux and generation of a persistent signal may account for the maintained elevation of [Ca2+]i.

Interleukin-1 stimulates phosphatidic acid-mediated phospholipase D activity in human mesangial cells

1994 ◽  
Vol 266 (4) ◽  
pp. C1093-C1104 ◽  
Author(s):  
S. L. Bursten ◽  
W. E. Harris
Keyword(s):  
Phosphatidic Acid ◽  
Phospholipase D ◽  
Mesangial Cells ◽  
Interleukin 1 ◽  
Type I ◽  
Human Mesangial Cells ◽  
Low Concentrations ◽  
Phosphatidate Phosphohydrolase ◽  
High Concentrations ◽  
Stimulation Of

Previous studies suggest that signal transduction mediated by interleukin-1 (IL-1), acting through an IL-1 receptor type found on T-cells and mesangial cells, may use phosphatidylethanolamine (PE) as a signaling molecule. Evidence presented here indicates that stimulation of human mesangial cells by IL-1 results in activation of a phospholipase D (PLD) that hydrolyzes PE to phosphatidic acid (PA). PLD acts on a subfraction of PE enriched in 1-o-alkyl and 1-o-alkenyl, sn-2-unsaturated species, generating a unique PA subspecies 30-120 s after stimulation. This PA species is subsequently converted to diradylglycerols by phosphatidate phosphohydrolase. The PE-directed PLD activity is abolished by antibodies against the IL-1 type I receptor and against IL-1. This specific PLD activity is also stimulated by low concentrations of 1,2-sn-dilinoleoyl PA, but not by high concentrations of 1-palmitoyl or 1-oleoyl lyso-PA. Blockade of PLD activation by IL-1 antibodies or antibody against the IL-1 receptor is bypassed by stimulation of human mesangial cells with 1,2-sn-dilinoleoyl PA. A novel system of signal cytokine mediation through PA self-amplification is indicated.

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