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:2.7.10.2 (focal adhesion kinase)
44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We studied the effects on plasma levels of coagulation and fibrinolysis factors of two currently used "sub-50" oral contraceptive preparations (OCs), one containing 750 micrograms lynestrenol and 37.5 micrograms ethinyl estradiol (LYN-EE) and the other containing 150 micrograms levonorgestrel and 30 micrograms ethinyl estradiol (LNG-EE), in groups of about 25 women aged 21 +/- 2 years. After 3 months, plasminogen levels increased in the two experimental groups (LYN-EE and LNG-EE), by 40% and 32%, respectively. This change was positively correlated with changes in ceruloplasmin levels, indicating that an estrogenic effect might be involved. Histidine-rich glycoprotein concentration decreased by 26% and 16%, respectively. Tissue-type plasminogen activator (t-PA) activity increased by 260% and 167%; t-PA antigen decreased by 12% and 18%, and t-PA inhibitor activity decreased by 31% and 32%, respectively. In the coagulation system, in both groups factor XII increased by 47% and 34%, respectively. The main inhibitor of factor XII, C1-inactivator, decreased slightly, but this was significant only in the LNG-EE group. The von Willebrand factor antigen fell by 8% and 9%, whereas factor VIII activity did not change. Antithrombin III antigen decreased by 14% in both groups. Factor IX activity increased by 15% and 21%. The difference in hormonal effects of both preparations was reflected by the increases in sex hormone binding globulin (by 130% and 21%) and ceruloplasmin (by 98% and 51%), indicating that LYN-EE had a more estrogenic potency than LNG-EE. In a control group of 25 matched subjects, who were observed simultaneously, we found no significant changes.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Effects of two low-dose oral contraceptives on circulating components of the coagulation and fibrinolytic systems. 310 29

Von Willebrand's disease (vWD), one of the most frequent hereditary bleeding disorders, is associated with a deficiency or a defective structure of von Willebrand factor (vWF). The defect is transmitted by autosomal inheritance. vWF is a plasma glycoprotein which mediates platelet adherence to the subendothelium of the injured blood vessel and is thus indispensable for primary hemostasis. vWF is composed of identical subunits linked together by disulfide bridges. Each subunit contains binding site(s) for factor VIII, collagen and platelets. The highly polymeric vWF factor is the largest known plasma protein. Functional activity is preferentially associated with the largest multimeric forms of vWF. Binding of vWF onto collagen fibrils activates its platelet binding sites, which are apparently not accessible in circulating vWF. Aggregation of washed fixed platelets in the presence of ristocetin or collagen is used for assessment of vWF's biological activity. Important additional information for the diagnosis of vWD is provided by the antigen assay and by electrophoretic analysis of the multimeric pattern of vWF. In type I vWD, all polymeric forms of vWF are reduced in the same proportion, while only the largest, i.e. the most active, multimers are deficient in type II vWD. The most severe type III vWD is characterized by an undetectable concentration of vWF. DDAVP corrects the prolonged bleeding time in patients with mild forms of vWD. Cryoprecipitate or "virus inactivated" factor VIII concentrates are employed for substitution therapy. Our results suggest that the newly introduced "virus inactivated" factor VIII concentrate from SRK is likewise suitable for this purpose.
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PMID:[Von Willebrand's disease]. 312 18

The adhesion of platelets to sites of vascular injury is critically dependent on the binding of subendothelial bound von Willebrand factor (vWf) to the platelet surface glycoprotein complexes, GP Ib-V-IX and GP IIb-IIIa (integrin alphaIIbbeta3). There is growing evidence that the binding of vWf to these receptors is not only essential for stable platelet adhesion but is also important for the transduction of activation signals required for changes in platelet morphology, granule secretion, and platelet aggregation. In this study we have investigated signaling events induced by vWf binding to GP Ib-V-IX in both spreading and aggregated platelets. The adhesion of platelets to vWf resulted in dramatic actin filament reorganization, as assessed by immunofluorescence with fluorescein isothiocyanate-conjugated phalloidin, and the cytoskeletal recruitment of various structural proteins (talin and integrin alphaIIbbeta3) and signaling enzymes (pp60c-src, focal adhesion kinase (FAK), phosphatidylinositol 3-kinase (PI 3-kinase), and protein-tyrosine phosphatase (PTP)-1B). Time course experiments in both spreading and aggregated platelets revealed that talin, FAK, and PTP-1B were proteolyzed after translocation to the cytoskeleton. The proteolysis of these proteins was dependent on the presence of extracellular calcium and was specifically inhibited by pretreating platelets with the membrane-permeable calpain inhibitors calpeptin, E64d, and MDL 28,170, but not with the membrane-impermeable inhibitors leupeptin, E64, and calpastatin. The cytoskeletal translocation of signaling enzymes in vWf-stimulated platelets was abolished by pretreating platelets with an anti-GP Ib-V-IX antibody but was unaffected by blocking ligand binding to integrin alphaIIbbeta3. In contrast, calpain activation in vWf-stimulated platelets required ligand binding to both GP Ib-V-IX and integrin alphaIIbbeta3. The activation of calpain in both spreading and aggregated platelets resulted in a substantial decrease in the level of tyrosine phosphorylation of multiple platelet proteins and was associated with a 50-80% reduction in the amount of cytoskeletal associated talin, integrin alphaIIbbeta3, PI 3-kinase, FAK, pp60(c-)src, and PTP-1B. These studies suggest a potentially important role for calpain in regulating the formation and/or stability of cytoskeletal signaling complexes in vWf-stimulated platelets. Furthermore, they demonstrate distinct roles for GP Ib-V-IX and integrin alphaIIbbeta3 in vWf-induced signal transduction.
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PMID:Calpain regulation of cytoskeletal signaling complexes in von Willebrand factor-stimulated platelets. Distinct roles for glycoprotein Ib-V-IX and glycoprotein IIb-IIIa (integrin alphaIIbbeta3) in von Willebrand factor-induced signal transduction. 926 16

Integrin alphaIIb beta3 mediates platelet aggregation and "outside-in" signaling. It is regulated by changes in receptor conformation and affinity and/or by lateral diffusion and receptor clustering. To document the relative contributions of conformation and clustering to alphaIIb beta3 function, alphaIIb was fused at its cytoplasmic tail to one or two FKBP12 repeats (FKBP). These modified alphaIIb subunits were expressed with beta3 in CHO cells, and the heterodimers could be clustered into morphologically detectable oligomers upon addition of AP1510, a membrane-permeable, bivalent FKBP ligand. Integrin clustering by AP1510 caused binding of fibrinogen and a multivalent (but not monovalent) fibrinogen-mimetic antibody. However, ligand binding due to clustering was only 25-50% of that observed when alphaIIb beta3 affinity was increased by an activating antibody or an activating mutation. The effects of integrin clustering and affinity modulation were additive, and clustering promoted irreversible ligand binding. Clustering of alphaIIb beta3 also promoted cell adhesion to fibrinogen or von Willebrand factor, but not as effectively as affinity modulation. However, clustering was sufficient to trigger fibrinogen-independent tyrosine phosphorylation of pp72(Syk) and fibrinogen-dependent phosphorylation of pp125(FAK), even in non-adherent cells. Thus, receptor clustering and affinity modulation play complementary roles in alphaIIb beta3 function. Affinity modulation is the predominant regulator of ligand binding and cell adhesion, but clustering increases these responses further and triggers protein tyrosine phosphorylation, even in the absence of affinity modulation. Both affinity modulation and clustering may be needed for optimal function of alphaIIb beta3 in platelets.
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PMID:Complementary roles for receptor clustering and conformational change in the adhesive and signaling functions of integrin alphaIIb beta3. 964 59

Two minisatellite (D1S80, D17S5) and 10 microsatellite (D2S1328, TPO, D3S1358, D9S926, D11S2010, THO1, VWF, FES, D16S310, and D18S848) polymorphic loci were analyzed in 5 Greek population groups (eastern Macedonia, central Macedonia, Thessaly, Epirus, and Greeks from Asia Minor) using the polymerase chain reaction. The genotypes at these loci conformed to Hardy-Weinberg equilibrium, and pairwise comparisons between them were in agreement with the expectation of independence between loci. This along with the low values of the coefficient of gene differentiation (GST) and the high heterozygosity levels of all loci allows the use of allele frequency data from the 12 hypervariable DNA markers for medicolegal casework in the Greek population groups studied. The small genetic distances indicate a genetic affinity among the 5 population samples. However, a few markers seem to allow some discrimination among the groups. No significant differences with other European populations were found for the loci studied.
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PMID:Genetic studies in 5 Greek population samples using 12 highly polymorphic DNA loci. 997 96

Proline-rich tyrosine kinase 2 (Pyk2) (also known as RAFTK, CAKbeta or CADTK) has been identified as a member of the focal adhesion kinase (FAK) family of protein-tyrosine kinases and it has been suggested that the mode of Pyk2 activation is distinct from that of FAK. In the present study we investigated the mode of Pyk2 activation in human platelets. When platelets were stimulated with thrombin, Pyk2, as well as FAK, was markedly tyrosine-phosphorylated, in a manner mostly dependent on alphaIIbbeta3 integrin-mediated aggregation. The residual Pyk2 tyrosine phosphorylation observed in the absence of platelet aggregation was completely abolished by pretreatment with BAPTA/AM [bis-(o-aminophenoxy)ethane-N,N,N',N'-tetra-acetic acid acetoxymethyl ester]. The Pyk2 phosphorylation was inhibited by protein kinase C (PKC) inhibitors at concentrations that inhibited platelet aggregation. In contrast, direct activation of PKC with the active phorbol ester PMA induced the tyrosine phosphorylation of Pyk2 and FAK but only when platelets were fully aggregated with the exogenous addition of fibrinogen (the ligand for alphaIIbbeta3 integrin). Furthermore, PMA-induced Pyk2 (and FAK) tyrosine phosphorylation was also observed when platelets adhered to immobilized fibrinogen. The activation of the von Willebrand factor (vWF)--glycoprotein Ib pathway with botrocetin together with vWF failed to induce Pyk2 (and FAK) tyrosine phosphorylation. Most Pyk2 and FAK was present in the cytosol and membrane skeleton fractions in unstimulated platelets. When platelets were stimulated with thrombin, both Pyk2 and FAK were translocated to the cytoskeleton in an aggregation-dependent manner. In immunoprecipitation studies, Pyk2, as well as FAK, seemed to associate with Shc through Grb2. With the use of glutathione S-transferase fusion proteins containing Shc-SH2, Grb2-SH2, and Grb2 N-terminal and C-terminal SH3 domains, it was implied that the proline-rich region of Pyk2 (and FAK) binds to the N-terminal SH3 domain of Grb2 and that the phosphotyrosine residue of Shc binds to the SH2 domain of Grb2. Although Pyk2 and FAK have been reported to be differentially regulated in many cell types, our results suggest that, in human platelets, the mode of Pyk2 activation is mostly similar to that of FAK, in terms of alphaIIbbeta3 integrin-dependent and PKC-dependent tyrosine phosphorylation. Furthermore, Pyk2, as well as FAK, might have one or more important roles in post-aggregation tyrosine phosphorylation events, in association with the cytoskeleton and through interaction with adapter proteins including Grb2 and Shc.
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PMID:Involvement of proline-rich tyrosine kinase 2 in platelet activation: tyrosine phosphorylation mostly dependent on alphaIIbbeta3 integrin and protein kinase C, translocation to the cytoskeleton and association with Shc through Grb2. 1074 87

The nonreceptor Bruton's tyrosine kinase (Btk) has been previously shown to associate physically and functionally with members of the protein kinase C (PKC) family of serine/threonine kinases in a variety of cell types. Here we show evidence for a novel interaction between Btk and PKCtheta; in platelets activated through the adhesion receptors GP Ib-V-IX and GP VI. Alboaggregin A, a snake venom component capable of activating both receptors in combination, leads to tyrosine phosphorylation of Btk downstream of Src family kinases. Inhibition of Btk by the selective antagonist LFM-A13 causes a reduction in calcium entry, although secretion of 5-hydroxytryptamine is potentiated. Btk is also phosphorylated on threonine residues in a PKC-dependent manner and associates with PKCtheta; upon platelet activation by either alboaggregin A or activation of GP Ib-V-IX alone by von Willebrand factor/ristocetin. PKCtheta; in turn becomes tyrosine-phosphorylated in a manner dependent upon Src family and Btk kinase activity. Inhibition of Btk activity by LFM-A13 leads to enhancement of PKCtheta; activity, whereas nonselective inhibition of PKC activity by bisindolylmaleimide I leads to reduction in Btk activity. We propose a reciprocal feedback interaction between Btk and PKCtheta; in platelets, in which PKCtheta; positively modulates activity of Btk, which in turn feeds back negatively upon PKCtheta;.
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PMID:Interaction of Bruton's tyrosine kinase and protein kinase Ctheta in platelets. Cross-talk between tyrosine and serine/threonine kinases. 1178 86

Stimulation of human platelets with von Willebrand factor (vWF) induces the rapid tyrosine phosphorylation of several proteins, but very little is known on the tyrosine kinases involved in this process. In the present work, we investigated and compared the activation of two related tyrosine kinases expressed in platelets: the proline-rich tyrosine kinase 2 (Pyk2) and the focal adhesion kinase (FAK). Both kinases were tyrosine phosphorylated upon vWF interaction with glycoprotein Ib-IX-V complex, but with different mechanisms. Tyrosine phosphorylation of FAK was totally dependent on thromboxane A2 production, and was inhibited by the integrin alphaIIbeta3 antagonist RGDS peptide. Moreover, chelation of intracellular calcium or inhibition of protein kinase C (PKC) totally blocked vWF-induced tyrosine phosphorylation of FAK, indicating that this event is downstream phospholipase A2 and phospholipase C activation. By contrast, tyrosine phosphorylation of Pyk2 was only partially reduced by aspirin and RGDS, and was not affected by either calcium chelation or PKC inhibition, suggesting that activation of this kinase does not require phospholipase-mediated signalling. Both FAK and Pyk2 translocated to the cytoskeleton upon vWF stimulation of human platelets by a mechanism depending on agonist-induced actin polymerisation. Prevention of cytoskeletal relocation of Pyk2 and FAK by cytochalasin D totally blocked vWF-induced tyrosine phosphorylation of both kinases. Finally, phosphorylation of Pyk2 induced by vWF, but not by thrombin, was inhibited by piceatannol, suggesting that this kinase lies downstream Syk. These results demonstrate that both Pyk2 and FAK are involved in platelet stimulation by vWF, but indicate that only Pyk2 may play a role in the early signal transduction events activated by ligand binding to glycoprotein Ib-IX-V.
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PMID:Proline-rich tyrosine kinase 2 and focal adhesion kinase are involved in different phases of platelet activation by vWF. 1191 84

After a vessel wall injury, platelets adhere to the subendothelium following a sequence of events: arrest of single platelets on the surface, progression to platelet spreading and final aggregation. Primary arrest of circulating platelets on subendothelial components occurs through platelet glycoprotein (GP) Ib and collagen receptors; then platelets spread and aggregate through a GPIIb-IIIa-dependent mechanism. A series of strategies were applied to analyse the tyrosine-phosphorylation mechanisms occurring at the different stages of platelet adhesion on subendothelial components under flow conditions, with special attention to primary arrest. To evaluate spread platelets, samples were exposed to acetylsalicylic acid, which blocks aggregate formation. To study single platelets in contact, a monoclonal antibody specific for GPIIb-IIIa was used to prevent platelet spreading and further aggregation. This experimental situation was also investigated using blood from two patients with Glanzmann's thrombasthenia (i.e. lacking GPIIb-IIIa). Results demonstrated that blockade of both spreading and aggregation results in significant changes in the tyrosine-phosphorylation patterns. Arrest of single platelets on collagen-rich surfaces resulted in phosphorylation of p125, identified as focal adhesion kinase (FAK), the 80/85 kDa doublet (cortactin), and p72, identified as Syk. Arrest of single platelets on von Willebrand factor as adhesive substrate showed that interaction through GPIb induces Syk phosphorylation, but not that of cortactin and FAK. Our data indicate that the initial arrest of platelets on subendothelial components involves Syk phosphorylation, which seems to be GPIb-dependent, and this is followed by activation and phosphorylation of cortactin and FAK. These processes seem to occur before GPIIb-IIIa becomes activated.
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PMID:Primary arrest of circulating platelets on collagen involves phosphorylation of Syk, cortactin and focal adhesion kinase: studies under flow conditions. 1198 77

The major platelet integrin alpha(IIb)beta(3), also known as the platelet glycoprotein (GP) IIb-IIIa complex, mediates platelet aggregation by serving as the receptor for fibrinogen and von Willebrand factor. In addition to its physiologic role, GPIIb-IIIa also bears a number of clinically important alloantigenic determinants. Previous studies have shown that disruption of the long-range Cys(5)-Cys(435) disulfide bond of the beta(3) subunit results in the production of isoforms that bind some, but not all, anti-Pl(A1) alloantibodies, suggesting that mutations in this so-called long-range disulfide bond can alter the conformation of GPIIIa. The purpose of this study was to examine the effects of either the Cys5Ala or Cys435Ala substitution of GPIIIa on the adhesive properties of the GPIIb-IIIa complex. We found that both Ala5GPIIIa and Ala435GPIIIa were capable of associating with GPIIb and were expressed normally on the cell surface when cotransfected into Chinese hamster ovary (CHO) cells. CHO cells expressing GPIIb-Ala5GPIIIa or GPIIb-Ala435IIIa bound well-characterized, conformationally sensitive ligand-induced binding site (LIBS) antibodies, and were capable of constitutively binding the fibrinogen-mimetic monoclonal antibodies Pl-55 and PAC-1, as well as soluble fibrinogen. Both GPIIb-Ala5IIIa- and GPIIb-Ala435IIIa-transfected CHO cells also bound more avidly to immobilized fibrinogen and were capable of mediating the tyrosine phosphorylation of pp125(FAK) on cell adhesion. These data are consistent with the notion that these regions of GPIIIa participate in the conformational change associated with receptor activation. Additionally, these studies may provide a molecular explanation for the previously reported ability of mild reducing agents to activate the GPIIb-IIIa complex and promote platelet aggregation.
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PMID:Disruption of the long-range GPIIIa Cys(5)-Cys(435) disulfide bond results in the production of constitutively active GPIIb-IIIa (alpha(IIb)beta(3)) integrin complexes. 1220 Mar 72


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