Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Serine esterase inhibitors (phenylmethanesulfonyl fluoride, 5-dimethylaminonaphthalene-1-sulfonyl (dansyl) fluoride, 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate, or p-nitrophenyl anthranilate) blocked the production of malonyldialdehyde by platelets induced with a variety of stimuli (including thrombin, trypsin, collagen, and A23187). These inhibitors did not block malonyldialdehyde production by platelets from exogenous arachidonic acid. Those inhibitors studied in greater detail (phenylmethanesulfonyl fluoride and 2-nitro-4-carboxyphenyl-N,N-diphenylcarbamate) were shown to inhibit the release of [1-14C]arachidonic acid from phosphatidylinositol and phosphatidylcholine in intact platelets but not the conversion of arachidonic acid to thromboxanes, prostaglandins, or hydroxyfatty acids. These inhibitors also blocked the stimulus-induced production of [32P]phosphatidic acid in intact platelets. Both arachidonic acid release from phosphatidylinositol and phosphatidic acid production have been reported to depend on the production of diglyceride by the action on the phosphatidylinositol-specific phospholipase C.f a phosphatidylinositol-specific phospholipase C. That enzyme in the soluble fraction from disrupted platelets was inhibited at concentrations of serine esterase inhibitors which block arachidonic acid release in intact platelets. These results indicate that serine esterase inhibitors block the stimulus-induced mobilization of arachidonic acid in platelets at least in part by their action o
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PMID:Serine esterase inhibitors block stimulus-induced mobilization of arachidonic acid and phosphatidylinositide-specific phospholipase C activity in platelets. 739 Oct 2

After binding to its receptor (uPAR), active cell-surface urokinase (uPA) is not internalized while the complex formed by uPA with plasminogen activator inhibitor type 1 (PAI-1) is internalized and degraded. Internalization and degradation require binding to uPAR and subsequently an interaction with the alpha 2-macroglobulin receptor (alpha 2-MR). To analyze the generality of this mechanism, we studied the internalization of uPA by recombinant protease nexin-1 (rPN-1), an inhibitor of thrombin, uPA, and plasmin. 125I-uPA.rPN-1 complexes bound specifically to uPAR; internalization occurred efficiently, and its time course was essentially the same as for uPA.PAI-1. Internalization required binding to uPAR since it could be blocked by the anti-uPAR monoclonal antibodies, by the uPAR antagonist amino-terminal fragment of uPA, and by the removal of uPAR by the treatment of cells with phosphatidylinositol-specific phospholipase C. As for uPA.PAI-1, the internalization of uPA.rPN-1 also required alpha 2-MR, since it could be inhibited by the 39-kDa alpha 2-macroglobulin receptor/low density lipoprotein receptor-associated protein, a ligand for the alpha 2-MR. Finally, we show by ligand blot analysis that the uPA.rPN-1 complex, like uPA.PAI-1 but unlike free uPA, bound specifically to both uPAR and alpha 2-MR.
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PMID:Protease nexin-1-urokinase complexes are internalized and degraded through a mechanism that requires both urokinase receptor and alpha 2-macroglobulin receptor. 802 43

Activation of human platelets with either the Escherichia coli alpha-haemolysin or thrombin, or with pharmacological agonists such as sodium fluoride (NaF), Ca-ionophore A23187 or phorbol myristate acetate (PMA), induced a similar pattern of serotonin release, unlike 12-hydroxyeicosatetraenoic acid (12-HETE) generation. In the presence of neomycin (0.1, 1, 10 mM), an inhibitor of the phosphatidylinositol-specific phospholipase C (PIP2-PLC), the E. coli alpha-haemolysin-induced 12-HETE generation was enhanced up to threefold in a dose-dependent manner. 12-HETE generation by NaF and thrombin was slightly inhibited at high neomycin concentrations (10 mM). Treatment of human platelets with E. coli alpha-haemolysin induced a different activation pattern of PIP2-PLC and phosphatidylinositol4-kinase (PI4-kinase), compared to NaF and thrombin. Haemolysin treatment resulted in a down-regulation of PIP2-PLC and PI4-kinase enzymatic activities by 20 +/- 9/40 +/- 8% compared to unstimulated cells; the decrease in enzymatic activities was observed within 2 min of stimulation and was still apparent after 60 min of stimulation. In NaF- and thrombin-stimulated platelets, dose- and time-dependent increases in PIP2-PLC (by up to 21 +/- 10%, 34 +/- 11%, respectively) and PI4-kinase (by up to 71 +/- 18, 54 +/- 14) activities were measured. Maximal enzymatic activities were reached after 5-20 min of stimulation (NaF, thrombin) followed by a decline to baseline levels (thrombin) or below baseline levels (NaF). Our results indicate that the phosphatidylinositolphosphate metabolism plays an important role in the regulation of 12-HETE release from human platelets by E. coli alpha-haemolysin.
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PMID:The role of the phosphatidylinositol turnover in 12-hydroxyeicosatetraenoic acid generation from human platelets by Escherichia coli alpha-haemolysin, thrombin and fluoride. 830 14

The translocation of protein kinase C (PKC) from the cytosolic to the particulate fraction in IIC9 fibroblasts has been studied to define the functions of 1,2-diacylglycerol (DAG) derived from the hydrolysis of phosphatidylinositol 4,5-bisphosphate (PIP2) and phosphatidylcholine (PC). alpha-Thrombin caused a biphasic change in DAG, with two peaks at 15-60 s and 5-15 min, derived from PIP2 and PC, respectively, while platelet-derived growth factor (PDGF) induced a monophasic DAG increase from PC at 5-15 min. alpha-Thrombin also induced a rapid, but transient, increase of inositol 1,4,5-trisphosphate and cytosolic Ca2+, whereas PDGF did not. Three PKC isozymes, alpha, epsilon, and zeta, were identified by Western blotting in IIC9 cells and were mainly localized in the cytosol. A fraction of cytosolic PKC alpha was rapidly translocated by alpha-thrombin at 15 s, but its membrane association was lost within 1 min. PKC epsilon was also rapidly translocated; however, its membrane association was sustained for almost 60 min. PKC zeta was not translocated by alpha-thrombin or phorbol 12-myristate 13-acetate. PDGF translocated PKC epsilon at 5 min but had little effect at 15 s and did not translocate PKC alpha or zeta. Incubation with Bacillus cereus PC- or phosphatidylinositol-specific phospholipase C, which increased DAG but not phosphatidic acid, stimulated translocation of PKC epsilon, but not PKC alpha or zeta. Addition of chelators to inhibit the rise in intracellular Ca2+ largely blocked PKC alpha translocation induced by alpha-thrombin but had no effect on PKC epsilon translocation. Addition of ionomycin allowed alpha-thrombin to induce PKC alpha translocation at 5 min. PKC alpha translocation was mimicked by 1,2-dioctanoylglycerol plus ionomycin, but not by either alone. On the other hand, PKC epsilon was translocated by the DAG alone. These results support the conclusion that PIP2 hydrolysis activates both PKC alpha and epsilon at 15 s, whereas PC hydrolysis activates only PKC epsilon at 5 min. The differential activation at 5 min can be attributed to the failure of PC hydrolysis to increase Ca2+ and not to a difference in the molecular species of DAG derived from the phospholipids.
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PMID:Differential translocation of protein kinase C isozymes by thrombin and platelet-derived growth factor. A possible function for phosphatidylcholine-derived diacylglycerol. 848 6

We determined whether activation of phosphatidylinositol-specific phospholipase C (PI-PLC) and a subsequent increase in cytosolic calcium concentration ([Ca2+]i) was an obligatory signaling event mediating the increase in transendothelial permeability induced by bradykinin (BK) and alpha-thrombin (alpha-T). Both BK and alpha-T (each at a concentration range of 0.01-1 microM) caused dose-dependent increases in transendothelial 125I-albumin permeability in cultured bovine pulmonary artery endothelial cell monolayers. Both agonists also produced a rise in inositol (1,4,5)-trisphosphate [Ins(1,4,5)P3] by 10 sec that was followed by a prolonged increase in [Ca2+]i. Pretreatment of endothelial cells with the PLC inhibitor, 1-(6-((17 beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1 H-pyrrole-2,5-dion [(U73122) at 10 microM for 15 min], prevented the increases in Ins(1,4,5)P3 and [Ca2+]i induced by both BK and alpha-T. However, inhibition of PLC with U73122 or another PLC inhibitor, neomycin, did not prevent the increase in endothelial permeability induced by either agonist. In contrast, depletion of cellular protein kinase C (PKC) with phorbol-12-myristate 13-acetate (0.01 microM for 20 hr) increased both BK- and alpha-T-induced phosphoinositide turnover but inhibited the agonist-induced increase in permeability. A PKC inhibitor, staurosporine (5 microM) likewise inhibited the BK-induced increase in endothelial cell permeability to albumin. We conclude that increases in endothelial permeability induced by the inflammatory mediators, BK and thrombin, can occur independently of PLC activation and increased [Ca2+]i but that a PKC-dependent pathway is required for the permeability response.
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PMID:Bradykinin- and thrombin-induced increases in endothelial permeability occur independently of phospholipase C but require protein kinase C activation. 936 52

Phospholipase Cepsilon (PLCepsilon) is one of the newest members of the phosphatidylinositol-specific phospholipase C (PLC) family. Previous studies have suggested that G-protein-coupled receptors (GPCRs) stimulate phosphoinositide (PI) hydrolysis by activating PLCbeta isoforms through G(q) family G proteins and Gbetagamma subunits. Using RNA interference to knock down PLC isoforms, we demonstrate that the GPCR agonists endothelin (ET-1), lysophosphatidic acid (LPA), and thrombin, acting through endogenous receptors, couple to both endogenous PLCepsilon and the PLCbeta isoform, PLCbeta3, in Rat-1 fibroblasts. Examination of the temporal activation of these PLC isoforms, however, reveals agonist- and isoform-specific profiles. PLCbeta3 is activated acutely within the first minute of ET-1, LPA, or thrombin stimulation but does not contribute to sustained PI hydrolysis induced by LPA or thrombin and accounts for only part of ET-1 sustained stimulation. PLCepsilon, on the other hand, predominantly accounts for sustained PI hydrolysis. Consistent with this observation, reconstitution of PLCepsilon in knockdown cells dose-dependently increases sustained, but not acute, agonist-stimulated PI hydrolysis. Furthermore, combined knockdown of both PLCepsilon and PLCbeta3 additively inhibits PI hydrolysis, suggesting independent regulation of each isoform. Importantly, ubiquitination of inositol 1,4,5-trisphosphate receptors correlates with sustained, but not acute, activation of PLCepsilon or PLCbeta3. In conclusion, GPCR agonists ET-1, LPA, and thrombin activate endogenous PLCepsilon and PLCbeta3 in Rat-1 fibroblasts. Activation of these PLC isoforms displays agonist-specific temporal profiles; however, PLCbeta3 is predominantly involved in acute and PLCepsilon in sustained PI hydrolysis.
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PMID:G-protein-coupled receptor agonists activate endogenous phospholipase Cepsilon and phospholipase Cbeta3 in a temporally distinct manner. 1631 22


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