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

---

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

Activation of human platelets by cross-linking of the platelet low-affinity IgG receptor, the Fc gamma receptor IIA (Fc gamma-RIIA), or by collagen is associated with rapid phosphorylation on tyrosine of the non-receptor tyrosine kinase syk. Phosphorylation is still observed, albeit sometimes reduced, in the presence of a combination of a protein kinase C inhibitor, Ro 31-8220, and the intracellular calcium chelator, BAPTA-AM, demonstrating independence from phosphoinositide-specific phospholipase C (PLC) activity. In contrast, the combination of Ro 31-8220 and BAPTA-AM completely inhibits phosphorylation of syk in thrombin-stimulated platelets. Phosphorylation of syk increases its autophosphorylation activity measured in a kinase assay performed on syk immunoprecipitates. Fc gamma-RIIA also undergoes phosphorylation in syk immunoprecipitates from platelets activated by cross-linking of Fc gamma-RIIA but not by collagen, suggesting that it associates with the kinase. Consistent with this, tyrosine-phosphorylated Fc gamma-RIIA is precipitated by a glutathione S-transferase (GST) fusion protein containing the tandem src homology (SH2) domains of syk from Fc gamma-RIIA- but not collagen-activated cells. Two uncharacterized tyrosine-phosphorylated proteins of 40 and 65 kDa are uniquely precipitated by a GST fusion protein containing the tandem syk-SH2 domains in collagen-stimulated platelets. A peptide based on the antigen recognition activation motif (ARAM) of Fc gamma-RIIA, and phosphorylated on the two tyrosine residues found within this region, selectively binds syk from lysates of resting platelets; this interaction is not seen with a non-phosphorylated peptide. Kinase assays on Fc gamma-RIIA immunoprecipitates reveal the constitutive association of an unidentified kinase activity in resting cells which phosphorylates a 67 kDa protein. Syk is not detected in Fc gamma-RIIA immunoprecipitates from resting cells but associates with the receptor following activation and, together with Fc gamma-RIIA, is phosphorylated in the kinase assay in vitro. These results demonstrate that syk is activated by Fc gamma-RIIA cross-linking and collagen, independent of PLC, suggesting that it may have an important role in the early events associated with platelet activation. The association of syk with Fc gamma-RIIA appears to be mediated through the tandem SH2 domains in syk and the ARAM motif of Fc gamma-RIIA. A similar interaction may underlie the response to collagen, suggesting that its signalling receptor contains an ARAM motif.
...
PMID:Syk interacts with tyrosine-phosphorylated proteins in human platelets activated by collagen and cross-linking of the Fc gamma-IIA receptor. 748 83

The interaction between von Willebrand factor (vWF) and glycoprotein Ib (GPIb) induced by ristocetin or botrocetin resulted in associated platelet aggregation, protein tyrosine phosphorylation (PTP) of a 64 kDa protein, as detected by a monoclonal antibody against phosphotyrosine (PY-20), and intracellular Ca2+ elevation that is largely dependent upon Ca2+ influx in human platelets. It is of interest that 75-80, 97 and 125 kDa proteins which are strongly tyrosine-phosphorylated in platelet activation induced by thrombin and other agonists were not detected. Neither vWF nor a coaggregating agent (ristocetin or botrocetin) alone induced aggregation, [Ca2+]i elevation or the 64 kDa PTP. NMC-4, an antibody which inhibits both ristocetin- or botrocetin-induced vWF binding to GPIb, abolished the appearance of the 64 kDa PTP as well as other responses, suggesting that it is specifically induced by the GPIb-vWF interaction. Aspirin, or ONO-3708, a competitive inhibitor of thromboxane A2, did not modify the 64 kDa PTP, while [Ca2+]i elevation was moderately suppressed. Depletion of extracellular Ca2+ or RGD peptides suppressed neither the 64 kDa PTP nor aggregation. H-7, a protein kinase C inhibitor, did not inhibit the 64 kDa PTP, while staurosporine, a potent protein kinase inhibitor, inhibited the 64 kDa PTP and Ca2+ influx, but not aggregation, in a dose-dependent manner. It is suggested that the 64 kDa PTP is associated with platelet aggregation induced by the interaction between GPIb and vWF.
...
PMID:Protein tyrosine phosphorylation in human platelets induced by interaction between glycoprotein Ib and von Willebrand factor. 753 5

1. Endothelin-1 has anti-aggregatory properties, but the mechanism underlying this inhibitory action is unknown. This in vitro study investigates effects of endothelin-1 on thrombin-stimulated aggregation and intracellular free calcium concentration in human platelets and assesses the role of protein kinase C in the interactions between endothelin-1 and thrombin. Aggregation was measured turbidometrically and the intracellular free calcium concentration was determined with the fluorescent indicator fura 2-acetoxymethyl ester. 2. Endothelin-1 at concentrations from 10(-11) to 10(-6) mol/l had no effect on platelet aggregation or intracellular free calcium concentration but inhibited in a dose-dependent manner aggregation induced by 0.05 unit/ml thrombin (pD2 for inhibition by endothelin = 8.1 +/- 0.12). 3. Endothelin-1 at 10(-9) mol/l significantly decreased (P < 0.01) thrombin-stimulated aggregation from 81.4 +/- 1.5% (in the absence of endothelin-1) to 53.5 +/- 1.1% (in the presence of endothelin) and thrombin-stimulated intracellular free calcium concentration from 179 +/- 1.7 nmol/l to 140 +/- 1.8 nmol/l. 4. Preincubation of platelets with 10(-7) mol/l staurosporine (protein kinase C inhibitor), calphostin C (highly selective protein kinase C inhibitor) or 5-(N,N-hexamethylene) amiloride (highly selective Na(+)-H+ exchange blocker) significantly inhibited (P < 0.01) thrombin-stimulated platelet responses and suppressed the inhibitory effect of endothelin-1 on thrombin-induced aggregation and intracellular free calcium concentration.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Role of protein kinase C in the anti-aggregatory effects of endothelin-1 on human platelets. 753 39

A potent, proteinaceous inducer of platelet aggregation designated as IVa, has been purified to homogeneity from Cerastes cerastes venom by molecular sieve and ion exchange chromatography. It is composed of 2 subunits with total M(r) of 62,000 as shown by native gel chromatography and chemical cross-linking with disuccinimidyl suberate. It is not clear at the present time whether both subunits are identical gene products, however, both have identical N-terminal sequences for the first 15 amino acids. The protein has a pI above 9.6. IVa (0.1 micrograms/ml) could aggregate platelets up to 80% and was inhibited by p-APMSF, leupeptin, iodoacetamide, protein kinase C inhibitor, phosphatase inhibitor, ATP and PGE1, while it was insensitive to acetylsalicylic acid, ADP scavenger system, protein kinase A inhibitor and hirudin. Protein IVa is a serine proteinase with thrombin-like activity as it hydrolysed thrombin chromogenic substrate CBS 34.47, its aggregatory activity was partially inhibited by monoclonal antibodies against GPIb and the thrombin receptor, as was the thrombin, and its ability to induce intracellular Ca2+ release was blocked by pretreating platelets with thrombin. Unlike thrombin, the IVa protein showed very weak coagulant activity as indicated by plasma recalcification time and fibrinogen clotting time although it could hydrolyse fibrinogen alpha-chains.
...
PMID:Characterization of a potent platelet aggregation inducer from Cerastes cerastes (Egyptian sand viper) venom. 761 60

The goal of the present studies was to determine whether anion fluxes are involved in thrombin- and histamine-activated signal transduction pathways in human umbilical vein endothelial cells (HUVECs). 125Iodine (125I) efflux techniques were used to test the sensitivity of anion fluxes to increases in [Ca2+]i and activation of protein kinase C. HUVECs exhibited constant 125I efflux rates under basal conditions. Administration of thrombin or histamine stimulated an increase in 125I efflux rates which returned to control values after approximately 1-2 min. Since both agonists stimulate increases in [Ca2+]i, we tested the hypothesis that 125I efflux was sensitive to changes in [Ca2+]i. When HUVECs were exposed to ionomycin or thapsigargin, the 125I efflux rate increased and remained elevated for several minutes. In subsequent experiments, HUVECs were incubated with the cell permanent Ca2+ chelator, 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-AM, to buffer changes in [Ca2+]i. This treatment reduced both basal and thrombin-stimulated 125I efflux. However, when Ca2+ was removed from the efflux buffer and replaced with EGTA, peak thrombin-stimulated 125I efflux remained unchanged. This anion efflux was also sensitive to activation of protein kinase C since phorbol 12-myristate 13-acetate and phorbol, 12,13-dibutyrate blunted thrombin-mediated increases in 125I efflux. Preincubation of HUVECs with protein kinase C inhibitor peptide [19-36] antagonized the phorbol ester-mediated decrease in thrombin-stimulated 125I efflux. We suggest that 125I efflux in HUVECs represents a Ca(2+)-sensitive anion conductance and that intracellular Ca2+ release, but not extracellular Ca2+ influx, is sufficient to initiate channel activity.
...
PMID:Calcium-mobilizing agonists stimulate anion fluxes in cultured endothelial cells from human umbilical vein. 788 9

Heparin-binding epidermal growth factor-like growth factor (HB-EGF) is a potent smooth muscle cell mitogen of macrophage origin. To determine whether the HB-EGF gene is transcribed and regulated in mesangial cells, we measured HB-EGF mRNA levels in cultured rat mesangial cells by RNA blot analysis. A 2.5-kb HB-EGF mRNA was detected in unstimulated mesangial cells. The protein kinase C activator 12-O-tetradecanoylphorbol 13-acetate (TPA) increased HB-EGF mRNA levels by 15-fold in mesangial cells, and this induction of HB-EGF mRNA by TPA was both time- and dose-dependent. HB-EGF mRNA could also be stimulated by 10% fetal calf serum, ionomycin, thrombin, and endothelin-1. Staurosporine, a protein kinase C inhibitor, abolished the induction of HB-EGF mRNA by TPA and serum. To determine whether HB-EGF is mitogenic for mesangial cells, we transfected COS cells with HB-EGF expression plasmids. Culture medium from COS cells transfected with these plasmids increased 3H-thymidine incorporation in mesangial cells in a dose-dependent manner. To our knowledge, this is the first report that HB-EGF is expressed in renal cells. This inducible transcription of HB-EGF suggests that it may have an autocrine role in mesangial cell proliferation in kidney disease.
...
PMID:Induction of heparin-binding epidermal growth factor-like growth factor mRNA by protein kinase C activators. 799 89

Changes of intracellular activity of lysolecithin acyltransferase (LAT) during an interaction between endothelial cells (EC) and low-density lipoprotein (LDL) were investigated. Following an incubation of EC with LDL, endothelial LAT activity was assayed using [3H]lysophosphatidylcholine as the substrate. Stimulation of EC with either thrombin (0.01-1 U/ml) or Ca(2+)-ionophore A23187 (10(-10)-10(-7) M) dose- and time-dependently enhanced LAT activity in the presence of LDL (1 mg protein/ml), but no enhancement was observed in quiescent cells. Ionomycin together with 1-oleoyl-2-acetyl glycerol, a synthetic analog of diacylglycerols enhanced LAT activity in a similar degree to thrombin in the presence of LDL. Either staurosporine, a protein kinase C inhibitor or neomycin, a phospholipase C inhibitor completely blocked an increase of LAT activity in stimulated EC. Stimulation of EC with various agonists including 12-o-tetradecanoylphorbol-13-acetate, an activator of protein kinase C caused a marked increase in cellular uptake of LDL, and staurosporine inhibited the uptake. These results suggest that the transport of LDL into EC is facilitated by stimulation with thrombin and other agonists, and LDL subsequently activates intracellular LAT. Protein kinase C seems to mediate LDL uptake into EC. Intracellular regulatory roles of LDL in the presence of vasoactive substances were discussed.
...
PMID:Enhancement of lysolecithin acyltransferase activity by LDL in thrombin-stimulated porcine-cultured endothelial cells. 801 83

Recent studies on cloning of the thrombin receptor, which belongs to the family of G-protein-coupled receptors, suggest that thrombin cleaves a peptide from the extracellular N-terminus. A synthetic peptide of 14 amino acids corresponding to the sequence of the newly generated N-terminus was found to possess thrombin-like activity in several cells endowed with thrombin receptors. The relaxant and contractile effects of this thrombin receptor activating peptide (TRAP, Ser-Phe-Leu-Leu-Arg-Asn-Pro-Asn-Asp-Lys-Tyr-Glu-Pro-Phe) were investigated in porcine pulmonary arteries and compared with the action of thrombin. In PGF2 alpha-precontracted vessels with intact endothelium, TRAP (0.3-10 mumol/l) caused reversible transient and concentration-dependent relaxation which was absent after mechanical removal of the endothelium. Preincubation of the vessels with NG-nitro-L-arginine (200 mumol/l) markedly reduced the relaxation. The TRAP-induced relaxation was associated with an increase in cGMP in the arteries. In comparison to thrombin, TRAP (EC50: 0.8 mumol/l) was less potent by more than three orders of magnitude. In endothelium-denuded pulmonary arteries TRAP (1-20 mumol/l) caused a concentration-dependent contraction which was reversible after washout. The TRAP-induced contractile response was preceded by an increase in generation of inositol 1,4,5-triphosphate (IP3); the peak of IP3 accumulation was reached after 30 s. Compared with the contractile effect of thrombin, that of TRAP was weaker by three of magnitude. The vascular effect of TRAP was not inhibited by the thrombin inhibitors hirudin or heparin while the protein kinase C inhibitor staurosporine (0.1 mumol/l) preferentially inhibited the tonic phase of contraction.(ABSTRACT TRUNCATED AT 250 WORDS)
...
PMID:Relaxant and contractile responses of porcine pulmonary arteries to a thrombin receptor activating peptide (TRAP). 805 15

To define the regulation of chemoattractant receptors, epitope-tagged human formyl peptide and C5a receptor cDNAs (ET-FR and ET-C5aR) were stably expressed in rat basophilic leukemia, RBL-2H3 cells. An antibody (12CA5) specific to "ET" was used to immunoprecipitate ET-FR and ET-C5aR. fMLP and C5a caused time- and dose-dependent phosphorylation of their respective receptors. Phosphorylated ET-FR migrated as a single broad band between 50 and 70 kDa on SDS-polyacrylamide gel electrophoresis, whereas ET-C5aR exhibited both fast (39-45 kDa) and broadly (39-52 kDa) migrating forms. Fast form phosphorylation alone was observed at low concentrations of C5a (0.001-0.01 microM), or at early times (5-30 s) with a higher concentration of C5a (0.1 microM). Phorbol 12-myristate 13-acetate, thrombin, or antigen caused no phosphorylation of ET-FR but stimulated exclusively fast form phosphorylation of ET-C5aR. The protein kinase C inhibitor staurosporine did not inhibit phosphorylation of ET-FR but blocked the fast migrating component of phosphorylated ET-C5aR. Homologous desensitization correlated with ligand-induced phosphorylation of both receptors. Of note, ET-C5aR but not ET-FR underwent heterologous desensitization by antigen, phorbol 12-myristate 13-acetate, and thrombin. The data suggest that protein kinase C mediates heterologous phosphorylation and desensitization of C5aR but not FR, yet, both receptors are homologously desensitized by a staurosporine-resistant kinase.
...
PMID:Differences in phosphorylation of formylpeptide and C5a chemoattractant receptors correlate with differences in desensitization. 822 71

The protease, alpha-thrombin (alpha Th), affects myocardial cell contractility, a feature common among agents that induce hypertrophy. However, it is not known whether cardiac myocytes possess alpha Th receptors (alpha Th-R), or if long term treatment with alpha Th can enhance growth and gene expression. In the present study primary neonatal rat ventricular myocytes expressed a 3.6-kilobase mRNA species that hybridized with a rat alpha Th-R-specific probe. After 48 h, alpha Th induced hypertrophy, sarcomeric organization, and enhanced atrial natriuretic factor (ANF) expression, all of which were blocked by the alpha Th-selective protease inhibitor, D-Phe-Pro-Arg-chloromethyl ketone. The alpha Th-R agonist peptide, SFLLRNPND, was a potent activator of ANF expression, however, the non-agonist, FLLRNPND, was inactive. Transfection experiments showed the enhancement of ANF expression by alpha Th to be transcriptional. The abilities of alpha Th to induce myocyte hypertrophy and to augment ANF transcription and peptide production were inhibited by the protein kinase C inhibitor, chelerythrine, and by the tyrosine kinase inhibitor, tyrphostin. Thus, myocardial cell alpha Th-Rs are stimulated by the specific proteolytic actions of alpha Th, and pathways involving both protein kinase C and protein tyrosine kinases are required for subsequent hypertrophy and ANF expression. Further, these findings suggest a new role for extracellular proteases as regulators of myocardial cell gene expression and growth.
...
PMID:Myocardial alpha-thrombin receptor activation induces hypertrophy and increases atrial natriuretic factor gene expression. 839 12


<< Previous 1 2 3 4 5 6 7 Next >>

---