scholarly journals Endothelial inositol phosphate generation and prostacyclin production in response to G-protein activation by AlF4−

1989 ◽  
Vol 264 (3) ◽  
pp. 703-711 ◽  
Author(s):  
M K Magnússon ◽  
H Halldórsson ◽  
M Kjeld ◽  
G Thorgeirsson
Keyword(s):  
Arachidonic Acid ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
G Protein ◽  
Pertussis Toxin ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Ionophore A23187 ◽  
Kinase C

In order to elucidate the role of guanine-nucleotide-binding proteins (G-proteins) in endothelial prostacyclin (PGI2) production, human umbilical vein endothelial cells, prelabelled with either [3H]inositol or [3H]arachidonic acid, were stimulated with the non-specific G-protein activator aluminium fluoride (AlF4-). AlF4- caused a dose- and time-dependent generation of inositol phosphates, release of arachidonic acid and production of PGI2. The curves for the three events were similar. When the cells were stimulated in low extracellular calcium (60 nM), they released [3H]arachidonic acid and produced PGI2, but depleting the intracellular Ca2+ stores by pretreatment with the Ca2+ ionophore A23187 totally inhibited both events, although the cells still responded when extracellular Ca2+ was added. The Ca2+ ionophore did not inhibit the generation of inositol phosphates in cells maintained at low extracellular Ca2+. Pertussis toxin pretreatment (14 h) altered neither inositol phosphate nor PGI2 production in response to AlF4-. To investigate the functional role of the diacylglycerol/protein kinase C arm of the phosphoinositide system, the cells were pretreated with the protein kinase C activator 12-O-tetradecanoylphorbol 13-acetate (TPA) or the protein kinase C inhibitor 1-(5-isoquinolinylsulphonyl)-2-methylpiperazine (H7). TPA inhibited the AlF4(-)-induced inositol phosphate generation but stimulated both the release of arachidonic acid and the production of PGI2. H7 had opposite effects both on inositol phosphate generation and on PGI2 production. These results suggest that AlF4(-)-induced PGI2 production is mediated by a pertussis-toxin-insensitive G-protein which activates the phosphoinositide second messenger system. This production of PGI2 can be modulated by protein kinase C activation, both at the level of inositol phosphate generation and at the level of arachidonic acid release.

scholarly journals Diradylglycerols stimulate phospholipase A2 and subsequent exocytosis in ram spermatozoa. Evidence that the effect is not mediated via protein kinase C

1994 ◽  
Vol 297 (1) ◽  
pp. 225-232 ◽  
Author(s):  
E R S Roldan ◽  
C Fragio
Keyword(s):  
Arachidonic Acid ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phospholipase A2 ◽  
Kinase Inhibitor ◽  
Ionophore A23187 ◽  
Kinase C ◽  
Acrosomal Exocytosis ◽  
Phospholipase A2 Activity

We tested the hypothesis that the role of diacylglycerol (DAG) in sperm acrosomal exocytosis is related to the activation of phospholipase A2, and that this effect is not mediated via protein kinase C. Treatment of [14C]arachidonic acid-labelled ram spermatozoa with Ca2+ and the ionophore A23187 stimulated both liberation of arachidonic acid and acrosomal exocytosis. No changes in [14C]DAG or [14C]monoacylglycerol were found after stimulation of spermatozoa, thus suggesting that arachidonic acid may be released exclusively via phospholipase A2. An increase in the endogenous levels of diradylglycerols (DRGs), resulting from exposure either to the DAG kinase inhibitor R 59022 or to exogenous 1-oleoyl-2-acetyl-sn-glycerol or 1,2-dioctanoyl-sn-glycerol, led to an increase in both phospholipase A2 activity and exocytosis when cells were stimulated with A23187 and Ca2+. Addition of DRGs that do not stimulate protein kinase C(1,3-dioctanoylglycerol, 1-O-hexadecyl-2-acetyl-rac-glycerol) also resulted in an increase in phospholipase A2 activity and exocytosis. On the other hand, phorbol esters (phorbol 12,13-dibutyrate; phorbol 12-myristate 13-acetate) did not enhance enzyme activity or exocytosis. Finally, exposure to 1-O-hexadecyl-2-O-methyl-rac-glycerol, a compound known to inhibit protein kinase C, did not affect phospholipase A2 activity or acrosomal exocytosis. We therefore conclude that in spermatozoa the messenger role of DAG is related to the activation of phospholipase A2, which in turn would generate an array of metabolites directly or indirectly involved in bringing about exocytosis of the acrosome.

Mechanism of action of luteinizing hormone-releasing hormone in rat ovarian cells

1989 ◽  
Vol 67 (8) ◽  
pp. 962-967 ◽  
Author(s):  
Peter C. K. Leung ◽  
Jian Wang ◽  
Kenneth G. Baimbridge
Keyword(s):  
Signal Transduction ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Luteinizing Hormone ◽  
Inositol Phosphates ◽  
Ionophore A23187 ◽  
Luteinizing Hormone Releasing Hormone ◽  
Releasing Hormone ◽  
Ovarian Cells ◽  
Kinase C

The initial step in the signal transduction of luteinizing hormone-releasing hormone (LHRH) in rat ovarian cells is the hydrolysis of membrane polyphosphoinositides into inositol phosphates and 1,2-diacylglycerol. The former compounds, especially inositol 1,4,5-triphosphate, are known to cause the release of calcium from intracellular stores, while diacylglycerol is a potent activator of protein kinase C. LHRH causes a rapid and transient increase in intracellular concentrations of free calcium ions, by approximately 4.5-fold, in the majority of granulosa cells as assessed by fura-2 microspectrofluorimetry. Like LHRH, a calcium ionophore (A23187) and activators of protein kinase C attenuate the steroidogenic response of the cells to follicle-stimulating hormone, but enhance the formation of gonadotropin-induced prostaglandin formation. These results support the concept that stimulation of polyphosphoinositide hydrolysis is intimitely involved in the direct action of LHRH at the level of the ovary.Key words: signal transduction, calcium, protein kinase C, ovary, steroid hormones.

scholarly journals Hydrogen peroxide-induced c-fos expression is mediated by arachidonic acid release: role of protein kinase C

1993 ◽  
Vol 21 (5) ◽  
pp. 1259-1263 ◽  
Author(s):  
Gadiparthi N. Rao ◽  
Bernard Lassegue ◽  
Kathy K. Griendling ◽  
R. Wayne Alexander ◽  
Bradford C. Berk
Keyword(s):  
Hydrogen Peroxide ◽  
Arachidonic Acid ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Arachidonic Acid Release ◽  
Kinase C ◽  
Acid Release ◽  
Fos Expression

scholarly journals Inhibition of mitogen-activated protein kinase kinase does not impair primary activation of human platelets

1996 ◽  
Vol 318 (1) ◽  
pp. 207-212 ◽  
Author(s):  
Angelika G. BÖRSCH-HAUBOLD ◽  
Ruth M. KRAMER ◽  
Steve P WATSON
Keyword(s):  
Arachidonic Acid ◽  
Protein Kinase C ◽  
Protein Kinase ◽  
Phorbol Ester ◽  
Substrate Inhibition ◽  
Human Platelets ◽  
Mitogen Activated Protein Kinase ◽  
Ionophore A23187 ◽  
Kinase C ◽  
Mitogen Activated Protein

Mitogen-activated protein kinases (MAPKs), a family of protein serine/threonine kinases regulating cell growth and differentiation, are activated by a dual-specificity kinase through phosphorylation at threonine and tyrosine. We used a recently described selective inhibitor of the p42/p44mapk-activating enzyme, PD 98059 [2-(2´-amino-3´-methoxyphenyl)-oxanaphthalen-4-one], to investigate the role of the p42/p44mapk pathway in human platelets. PD 98059 inhibited p42/p44mapk activation in thrombin-, collagen- and phorbol ester-stimulated platelets, as determined from in-gel renaturation kinase assays, with an IC50 of approx. 5 µM (thrombin stimulation). It also prevented activation of MAPK kinase, which was measured in whole-cell lysates with glutathione S-transferase/p42mapk fusion protein (GST–MAPK) as substrate. Inhibition of p42/p44mapk did not affect platelet responses to thrombin or collagen such as aggregation, 5-hydroxytryptamine release and protein kinase C activation. In addition, PD 98059 did not interfere with release of arachidonic acid, a response mediated by cytosolic phospholipase A2 (cPLA2), or with cPLA2 phosphorylation. This suggests that platelet cPLA2 is not regulated by p42/p44mapk after stimulation with physiological agonists. In contrast, phorbol ester-induced phosphorylation of cPLA2 and potentiation of arachidonic acid release stimulated by Ca2+ ionophore A23187 were inhibited by PD 98059, indicating that p42/p44mapk phosphorylates cPLA2 after activation of protein kinase C by the non-physiological tumour promoter.

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