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.

Regulation of oxytocin secretion by the ovine corpus luteum: effect of activators of protein kinase C

1990 ◽  
Vol 124 (2) ◽  
pp. 225-232 ◽  
Author(s):  
J. J. Hirst ◽  
G. E. Rice ◽  
G. Jenkin ◽  
G. D. Thorburn
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Corpus Luteum ◽  
Phospholipase C ◽  
Tissue Slices ◽  
Kinase C ◽  
Luteal Tissue ◽  
Dose Dependent ◽  
Stimulation Of

ABSTRACT The effect of protein kinase C activation and dibutyryl cyclic AMP on oxytocin secretion by ovine luteal tissue slices was investigated. Several putative regulators of luteal oxytocin secretion were also examined. Oxytocin was secreted by luteal tissue slices at a basal rate of 234·4 ± 32·8 pmol/g per h (n = 24) during 60-min incubations.Activators of protein kinase C: phorbol 12,13-dibutyrate (n = 8), phorbol 12-myristate,13-acetate (n = 4) and 1,2-didecanoylglycerol (n = 5), caused a dose-dependent stimulation of oxytocin secretion in the presence of a calcium ionophore (A23187; 0·2 μmol/l). Phospholipase C (PLC; 50–250 units/l) also caused a dose-dependent stimulation of oxytocin secretion by luteal slices. Phospholipase C-stimulated oxytocin secretion was potentiated by the addition of an inhibitor of diacylglycerol kinase (R59 022; n = 4). These data suggest that the activation of protein kinase C has a role in the stimulation of luteal oxytocin secretion. The results are also consistent with the involvement of protein kinase C in PLC-stimulated oxytocin secretion. The cyclic AMP second messenger system does not appear to be involved in the control of oxytocin secretion by the corpus luteum. Journal of Endocrinology (1990) 124, 225–232

scholarly journals Targeting of Protein Kinase C-ϵ during Fcγ Receptor-dependent Phagocytosis Requires the ϵC1B Domain and Phospholipase C-γ1

2006 ◽  
Vol 17 (2) ◽  
pp. 799-813 ◽  
Author(s):  
Keylon L. Cheeseman ◽  
Takehiko Ueyama ◽  
Tanya M. Michaud ◽  
Kaori Kashiwagi ◽  
Demin Wang ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Phospholipase D ◽  
Fluorescent Protein ◽  
Wild Type ◽  
Fcγ Receptor ◽  
Kinase C ◽  
Green Fluorescent

Protein kinase C-ϵ (PKC-ϵ) translocates to phagosomes and promotes uptake of IgG-opsonized targets. To identify the regions responsible for this concentration, green fluorescent protein (GFP)-protein kinase C-ϵ mutants were tracked during phagocytosis and in response to exogenous lipids. Deletion of the diacylglycerol (DAG)-binding ϵC1 and ϵC1B domains, or the ϵC1B point mutant ϵC259G, decreased accumulation at phagosomes and membrane translocation in response to exogenous DAG. Quantitation of GFP revealed that ϵC259G, ϵC1, and ϵC1B accumulation at phagosomes was significantly less than that of intact PKC-ϵ. Also, the DAG antagonist 1-hexadecyl-2-acetyl glycerol (EI-150) blocked PKC-ϵ translocation. Thus, DAG binding to ϵC1B is necessary for PKC-ϵ translocation. The role of phospholipase D (PLD), phosphatidylinositol-specific phospholipase C (PI-PLC)-γ1, and PI-PLC-γ2 in PKC-ϵ accumulation was assessed. Although GFP-PLD2 localized to phagosomes and enhanced phagocytosis, PLD inhibition did not alter target ingestion or PKC-ϵ localization. In contrast, the PI-PLC inhibitor U73122 decreased both phagocytosis and PKC-ϵ accumulation. Although expression of PI-PLC-γ2 is higher than that of PI-PLC-γ1, PI-PLC-γ1 but not PI-PLC-γ2 consistently concentrated at phagosomes. Macrophages from PI-PLC-γ2-/-mice were similar to wild-type macrophages in their rate and extent of phagocytosis, their accumulation of PKC-ϵ at the phagosome, and their sensitivity to U73122. This implicates PI-PLC-γ1 as the enzyme that supports PKC-ϵ localization and phagocytosis. That PI-PLC-γ1 was transiently tyrosine phosphorylated in nascent phagosomes is consistent with this conclusion. Together, these results support a model in which PI-PLC-γ1 provides DAG that binds to ϵC1B, facilitating PKC-ϵ localization to phagosomes for efficient IgG-mediated phagocytosis.

Leptin Constrains Phospholipase C-Protein Kinase C-Induced Insulin Secretion via a Phosphatidylinositol 3-Kinase-Dependent Pathway

2003 ◽  
Vol 228 (2) ◽  
pp. 175-182 ◽  
Author(s):  
Joo-Won Lee ◽  
Andrew G. Swick ◽  
Dale R. Romsos
Keyword(s):  
Protein Kinase C ◽  
Insulin Secretion ◽  
Protein Kinase ◽  
Phospholipase C ◽  
Phosphatidylinositol 3 Kinase ◽  
C Protein ◽  
Kinase C ◽  
Pka Pathway ◽  
Stimulation Of ◽  
Phosphatidylinositol 3

Leptin-deficient Lepob/Lepob mice hypersecrete insulin in response to acetylcholine stimulation of the phospholipase C-protein kinase C (PLC-PKC) pathway, and leptin constrains this hypersecretion. Leptin has been reported to activate phosphatidylinositol 3-kinase (PI 3-K) and subsequently phosphodiesterase (PDE) to impair protein kinase A (PKA)-induced insulin secretion from cultured islets of neonatal rats. We determined if PKA-induced insulin secretion was also hyperresponsive in Islets from Lepob/Lepob mice, and if leptin impaired this pathway in islets from these mice. Additionally, the possible role for PI 3-K and PDE in leptin-induced control of acetylcholine-induced insulin secretion was examined. Stimulation of Insulin secretion with GLP-1, forskolin (an activator of adenylyl cyclase), or IBMX (an inhibitor of PDE) did not cause hypersecretion of insulin from islets of young Lepob/Lepob mice, and leptin did not inhibit GLP-1-induced insulin secretion from islets of these mice. Inhibition of PDE with IBMX also did not block leptin-induced inhibition of acetylcholine-mediated insulin secretion from islets of Lepob/Lepob mice. But, preincubation of islets with wortmannin, an Inhibitor of PI 3-K activity, blocked the ability of leptin to constrain acetylcholine-induced insulin secretion from islets of Lepob/Lepob mice. We conclude that the capacity of the PKA pathway to stimulate insulin secretion is not increased in islets from young Lepob/Lepob mice, and that leptin does not regulate this pathway in islets from mice. Leptin may stimulate PI 3-K to constrain PLC-PKC-induced insulin secretion from Islets of Lepob/Lepob mice.

Endothelin activation of phospholipase D: dual modulation by protein kinase C and Ca2+

1993 ◽  
Vol 264 (5) ◽  
pp. F845-F853
Author(s):  
M. M. Friedlaender ◽  
D. Jain ◽  
Z. Ahmed ◽  
D. Hart ◽  
R. L. Barnett ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase D ◽  
Intracellular Signaling ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Eta Receptor ◽  
Ca2 Channels ◽  
Stimulation Of

Previous work from this laboratory has identified an endothelin (ET) type A (ETA) receptor on cultured rat renal medullary interstitial cells (RMIC), coupled to phosphatidylinositol-specific phospholipase C (PI-PLC), dihydropyridine-insensitive receptor-operated Ca2+ channels, and phospholipase A2. The current studies explored a role for ET stimulation of phosphatidylcholine-specific phospholipase D (PC-PLD) in intracellular signaling of this cell type. ET stimulated PLD activation, as measured by phosphatidic acid (PA) or phosphatidylethanol (PEt) accumulation, in a time- and concentration-dependent manner. Inhibition of diacylglycerol (DAG) kinase by ethylene glycol dioctanoate or 6-(2)4-[(4-fluorophenyl)-phenylmethylene]-1-piperadinyl]ethy l-7-methyl-5H - thiaxolo-[3,2-alpyrimidin]-5-one (R 59022) failed to blunt PA accumulation, indicating that PLD, and not DAG, was the source of PA. Inhibition of PA phosphohydrolase (PAP) by propranolol increased late accumulation of PA, suggesting that the prevailing metabolic flow was in the direction of PA to DAG. Phorbol 12-myristate 13-acetate (PMA) augmented ET-evoked PEt accumulation, whereas downregulation of protein kinase C (PKC) obviated agonist-induced PEt production. PMA augmentation of PLD activity proceeded independent of cytosolic free Ca2+ concentration. Ca2+ derived from either intracellular or extracellular sources enhanced ET-related PEt accumulation but was without effect in PKC-downregulated cells. Collectively, these observations indicate that ET stimulates PLD production in RMIC. PKC is the major regulator of this process, with Ca2+ playing a secondary, modulatory role. In addition, these data suggest that PC-PLD is coupled to the ETA receptor.

scholarly journals Stimulation of tissue plasminogen activator production by retinoic acid: synergistic effect on protein kinase C-mediated activation

Blood ◽  
1992 ◽  
Vol 80 (4) ◽  
pp. 981-987 ◽  
Author(s):  
RD Medh ◽  
L Santell ◽  
EG Levin
Keyword(s):  
Endothelial Cells ◽  
Protein Kinase C ◽  
Retinoic Acid ◽  
Protein Kinase ◽  
Hela Cells ◽  
Additive Effect ◽  
Kinase C ◽  
Tissue Plasminogen ◽  
Camp Levels ◽  
Stimulation Of

Abstract Trans retinoic acid (t-RA) stimulated the production of tissue plasminogen activator (tPA) in HeLa-S3 and human umbilical vein endothelial cells (huvecs) in a dose-dependent manner with maximal release (four to five times control) at 40 nmol/L and 40 mumol/L, respectively. In endothelial cells, the stimulation of tPA production by phorbol 12-myristate 13-acetate (PMA) was potentiated 1.9-fold by 10 mumol/L t-RA, or 1.8 times the additive effect. In HeLa cells, total tPA secretion with 10 nmol/L PMA was increased from 43 ng/mL to 96 ng/mL by 40 nmol/L t-RA, which was two times the additive effect. Higher concentrations of t-RA (400 nmol/L) depressed tPA secretion by itself and also suppressed PMA-induced tPA production by 50%. Histamine and thrombin also synergized with t-RA. t-RA (40 nmol/L) and 10 micrograms/mL histamine or 10 U/mL thrombin combined to induce tPA production 3.4 and 1.3 times the additive effect in HeLa cells. Cyclic adenosine monophosphate (cAMP) levels were not significantly affected by 10 nmol/L to 10 mumol/L t-RA. Nor did 10 nmol/L PMA and 40 nmol/L t- RA together affect cAMP levels, suggesting that t-RA-mediated potentiation of PMA-induced tPA production occurred via a mechanism that was independent of cAMP levels. Downregulation of protein kinase C (PKC) by pretreatment of huvecs with 100 nmol/L PMA completely blocked a secondary response to PMA, but did not have a significant effect on t- RA induction. Pretreatment with 10 mumol/L t-RA, on the other hand, did not significantly affect a secondary stimulus by 100 nmol/L PMA, but completely suppressed a secondary stimulation by 10 mumol/L t-RA alone. These studies suggest that the mechanism mediating t-RA stimulation of tPA production interacts with the PKC pathway, resulting in synergism.

Phospholipase D: enzymology, mechanisms of regulation, and function

1997 ◽  
Vol 77 (2) ◽  
pp. 303-320 ◽  
Author(s):  
J. H. Exton
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Phosphatidic Acid ◽  
G Protein ◽  
Phospholipase D ◽  
Tyrosine Kinases ◽  
Rho Proteins ◽  
Kinase C

Phospholipase D exists in various forms that differ in their regulation but predominantly hydrolyze phosphatidylcholine. The Ca(2+)-dependent isozymes of protein kinase C regulate phospholipase D in vitro and play a major role in its control by growth factors and G protein-linked agonists in vivo. Recent studies have demonstrated that small G proteins of the ADP-ribosylation factor (ARF) and Rho families activate the enzyme in vitro, and evidence is accumulating that they also are involved in its control in vivo. Both types of G protein play important roles in cellular function, and the possible mechanisms by which they are activated by agonists are discussed. There is also emerging evidence of the control of phospholipase D and Rho proteins by soluble tyrosine kinases and novel serine/threonine kinases. The possible role of these kinases in agonist regulation of phospholipase D is discussed. The function of phospholipase D in cells is still poorly defined. Postulated roles of phosphatidic acid produced by phospholipase D action include the activation of Ca(2+)-independent isoforms of protein kinase C, the regulation of growth and the cytoskeleton in fibroblasts, and control of the respiratory burst in neutrophils. Another important function of phosphatidic acid is to act as a substrate for a specific phospholipase A2 to generate lysophosphatidic acid, which is becoming increasingly recognized as a major intercellular messenger. Finally, it is possible that the phospholipid changes induced in various cellular membranes by phospholipase D may per se play an important role in vesicle trafficking and other membrane-associated events.

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