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)

A number of cytokines, including basic fibroblast growth factor (bFGF), vascular endothelial growth factor (VEGF), oncostatin M (OSM), IL-6, and tumor necrosis factor alpha (TNF-alpha), have been postulated to have a role in the pathogenesis of Kaposi's sarcoma (KS). The proliferative effects of bFGF and OSM may be via their reported activation of the c-Jun NH2-terminal kinase (JNK) signaling pathway in KS cells. We now report that KS cells express a recently identified focal adhesion kinase termed RAFTK which appears in other cell systems to coordinate surface signals between cytokine and integrin receptors and the cytoskeleton as well as act downstream to modulate JNK activation. We also report that the tyrosine kinase receptor FLT-4, present on normal lymphatic endothelium, is robustly expressed in KS cells. Treatment of KS cells with VEGF-related protein (VRP), the ligand for the FLT-4 receptor, as well as with the cytokines bFGF, OSM, IL-6, VEGF, or TNF-alpha resulted in phosphorylation and activation of RAFTK. Following its activation, there was an enhanced association of RAFTK with the cytoskeletal protein paxillin. This association was mediated by the hydrophobic COOH-terminal domain of the kinase. Furthermore, JNK activity was increased in KS cells after VEGF or VRP stimulation. We postulate that in these tumor cells RAFTK may be activated by a diverse group of stimulatory cytokines and facilitate signal transduction to the cytoskeleton and downstream to the growth promoting JNK pathway.
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PMID:Cytokine signaling through the novel tyrosine kinase RAFTK in Kaposi's sarcoma cells. 912 25

Alveolar epithelial type II cells are the progenitor cells for restoring the alveolar epithelial barrier after acute lung injury. During repair of lung injury, the alveolar epithelial type II cells reepithelialize denuded air spaces, a process that involves breaking and reforming cell adhesions. A novel technique of mechanical separation of cultured alveolar epithelial cells from in vitro matrix was used to examine the intracellular signals that result when alveolar epithelial cell adhesions are broken. The results show that the tyrosine phosphorylation levels of focal adhesion kinase, paxillin, and pp60(src) decreased immediately after mechanical separation of the cells. Levels returned to nearly normal by 24 h after mechanical separation. Paxillin and pp60(scr) coprecipitated with focal adhesion kinase regardless of their phosphorylation state. Interestingly, the tyrosine phosphorylation level of the mitogen-activated protein kinase, p42(erk2), increased 15 min after mechanical separation. Preincubation of cell monolayers with phenylarsine oxide, a protein tyrosine phosphatase inhibitor, blocked the decrease in tyrosine phosphorylation levels of focal adhesion kinase, paxillin and pp60(src). Phenylarsine oxide incubation also prevented readhesion of mechanically separated cells at 24 h, but genistein, a tyrosine kinase inhibitor, had no effect. We conclude that protein tyrosine phosphatases are activated immediately after cultured alveolar epithelial cells are mechanically separated from in vitro matrix, and their activation is required for alveolar epithelial cell readhesion.
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PMID:Protein tyrosine phosphatases mediate cell readhesion in alveolar epithelial cells mechanically separated from in vitro matrix. 916 Aug 44

We and others have recently cloned a non-receptor, calcium-dependent tyrosine kinase (CADTK; also known as PYK2, CAKbeta, and RAFTK) that shares both overall domain structure and 45% amino acid identity with p125(FAK). We have studied the signaling, activation, and potential function of these related enzymes in GN4 rat liver epithelial cells that express CADTK and p125(FAK) at roughly similar levels. p125(FAK) is nearly fully tyrosine-phosphorylated in resting GN4 cells. In contrast, while CADTK is not tyrosine-autophosphorylated in untreated cells, angiotensin II increases CADTK Tyr(P) by 5-10-fold. With regard to signaling, CADTK activation is correlated with stimulation of c-Jun N-terminal kinase and p70(S6K) pathways but not with the stimulation of mitogen-activated protein kinase or p90(RSK). In this report we assessed the contribution of CADTK and p125(FAK) to tyrosine phosphorylation of focal contact proteins. In adherent GN4 cells, the constitutive activity of p125(FAK) was correlated with basal paxillin, tensin, and p130(CAS) tyrosine phosphorylation. A rapid increase in the tyrosine phosphorylation of each protein was detected after treatment with angiotensin II or other agonists that stimulate CADTK; the prolonged 3-4-fold increase in paxillin tyrosine phosphorylation was the most substantial change. In the WB cell line that expresses 3-fold less CADTK than GN4 cell line agonist-dependent paxillin tyrosine phosphorylation is similarly reduced. Immunoprecipitation of CADTK from GN4 cells revealed CADTK. paxillin complexes that persisted in 500 mM NaCl but not in 0.1% SDS cell lysis buffer. The complexes were largely independent of the tyrosine phosphorylation state of either protein. Surprisingly, we did not detect p125(FAK).paxillin complexes in immunoprecipitates using either of two p125(FAK) antibodies. When CADTK and p125(FAK) were transiently overexpressed in 293(T) cells, both enzymes associated with paxillin, but the avidity of CADTK appeared to be greater. In addition, in transfected 293(T) cells, complexes between CADTK and another potential substrate, p130(CAS), were detected. In summary, in GN4 rat liver epithelial cells stimulation of CADTK was highly correlated with paxillin tyrosine phosphorylation; in addition, CADTK but not p125(FAK) was complexed to paxillin at detectable levels. This suggests that agonist-dependent cytoskeletal changes in epithelial cells might proceed, in part, by CADTK-dependent mechanisms.
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PMID:Paxillin is tyrosine-phosphorylated by and preferentially associates with the calcium-dependent tyrosine kinase in rat liver epithelial cells. 916 70

Treatment of T47-D human breast carcinoma cells with recombinant prolactin (rhPRL) induced a concentration- and time-dependent increase in the phosphotyrosine content of JAK2. rhPRL also stimulated JAK2 tyrosine phosphorylation more weakly in three other breast carcinoma lines, MCF-7, ZR-75-1 and MDA-MB-231. Furthermore it stimulated tyrosine phosphorylation of two isoforms of the transcriptional activator STAT5, STAT5a and STAT5b. Surprisingly, rhPRL treatment of T47-D cells also stimulated tyrosine phosphorylation of focal adhesion kinase (FAK), as determined by immunoprecipitation with anti-phosphotyrosine antibody followed by immunoblotting with a specific FAK antibody. The effect of rhPRL was rapid and concentration-dependent, being maximal at 5 ng/ml. At rhPRL concentrations above 25 ng/ml, FAK tyrosine phosphorylation declined but remained above control levels at 100 ng/ml. rhPRL also stimulated paxillin tyrosine phosphorylation in T47-D cells with similar concentration- and time-dependence. In a second human breast carcinoma cell line, MCF-7, rhPRL produced very similar effects on FAK and paxillin tyrosine phosphorylation. These findings identify a new protein tyrosine kinase pathway in the action of the lactogenic hormone rhPRL and represent the first report that a hormone acting through a member of the haemopoietin receptor superfamily can regulate the FAK/paxillin pathway.
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PMID:Prolactin stimulates the JAK2 and focal adhesion kinase pathways in human breast carcinoma T47-D cells. 916 61

Assembly of a fibronectin (FN) matrix is a multistep process which influences a number of cellular functions including intracellular cytoskeletal organization and signaling responses. We have previously reported on a recombinant FN (recFN), FN delta III1-7, which differs from native FN in its rate of fibril formation. To determine the intracellular consequences of a delay in assembly, we compared the distribution of cytoskeletal proteins during the formation of native and recFN matrices by immunofluorescence at various time points. CHO alpha 5 cell cytoskeleton was reorganized in response to both native and recFN matrix formation. Assembly of native FN induced a rapid reorganization of actin into stress fibers and colocalization of alpha 5 beta 1 integrin, focal adhesion kinase (FAK), vinculin, and paxillin to regions of cell-matrix contact. alpha 5 beta 1 integrins and FAK are also clustered upon binding of FN delta III1-7 to cells but actin reorganization and focal adhesion formation are delayed and appear to be dependent on the formation of FN delta III1-7 fibrils. These results suggest that the structural framework of the matrix plays an important role in the ability of FN to initiate intracellular responses.
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PMID:Coordinated regulation of fibronectin fibril assembly and actin stress fiber formation. 917 3

Vascular endothelial growth factor (VEGF) stimulated the tyrosine phosphorylation of multiple components in confluent human umbilical vein endothelial cells (HUVECs) including bands of Mr 205,000, corresponding to the VEGF receptors Flt-1 and KDR, and Mr 145,000, 120,000, 97,000, and 65,000-70,000. VEGF caused a striking and transient increase in mitogen-activated protein (MAP) kinase activity and stimulated phospholipase C-gamma tyrosine phosphorylation, but it had no effect on phosphatidylinositol 3'-kinase activity. VEGF caused a marked increase in tyrosine phosphorylation of p125 focal adhesion kinase (p125(FAK)), which was both rapid and concentration-dependent. VEGF produced similar effects on p125(FAK) in the endothelial cell line ECV.304. VEGF stimulated tyrosine phosphorylation of the 68-kDa focal adhesion-associated component, paxillin, with similar kinetics and concentration dependence to that for p125(FAK). Thrombin and the phorbol ester, phorbol 12-myristate 13-acetate, also increased p125(FAK) tyrosine phosphorylation in HUVECs. The effect of VEGF on p125(FAK) tyrosine phosphorylation was completely inhibited by the actin filament-disrupting agent cytochalasin D and was partially inhibited by the protein kinase C inhibitor GF109203X. Inhibition of the MAP kinase pathway using a specific inhibitor of MAP kinase kinase had no effect on p125(FAK) tyrosine phosphorylation. VEGF stimulated migration and actin stress fiber formation in confluent HUVEC, and VEGF-induced p125(FAK)/paxillin tyrosine phosphorylation was accompanied by increased immunofluorescent staining of p125(FAK), paxillin, and phosphotyrosine in focal adhesions in confluent cultures of HUVECs. These findings identify p125(FAK) and paxillin as components in a VEGF-stimulated signaling pathway and suggest a novel mechanism for VEGF regulation of endothelial cell functions.
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PMID:Vascular endothelial growth factor stimulates tyrosine phosphorylation and recruitment to new focal adhesions of focal adhesion kinase and paxillin in endothelial cells. 918 76

Sphingosine 1-phosphate (SPP), a sphingolipid second messenger implicated in the mitogenic action of platelet-derived growth factor [Olivera, A. and Spiegel, S. (1993) Nature (London) 365, 557-560], induced rapid reorganization of the actin cytoskeleton resulting in stress-fibre formation. SPP also induced transient tyrosine phosphorylation of focal adhesion kinase (p125(FAK)), a cytosolic tyrosine kinase that localizes in focal adhesions, and of the cytoskeleton-associated protein paxillin. Exoenzyme C3 transferase, which ADP-ribosylates Rho (a Ras-related small GTP binding protein) on asparagine-41 and renders it biologically inactive, inhibited both stress-fibre formation and protein tyrosine phosphorylation induced by SPP. Thus Rho may be an upstream regulator of both stress-fibre formation and tyrosine phosphorylation of p125(FAK) and paxillin. Pretreatment with PMA, an activator of protein kinase C (PKC), inhibited the stimulation of stress-fibre formation induced by 1-oleoyl-lysophosphatidic acid (LPA) but not that by SPP. Similarly, PMA also decreased LPA-induced tyrosine phosphorylation of p125(FAK) and paxillin without abrogating the response to SPP. Thus PKC is involved in LPA- but not SPP-dependent signalling. The polyanionic drug suramin, a broad-specificity inhibitor of ligand-receptor interactions, did not inhibit either the mitogenic effect of SPP or its stimulation of tyrosine phosphorylation of p125(FAK). However, suramin markedly inhibited these responses induced by LPA. These results suggest that in contrast with LPA, SPP may be acting intracellularly in Swiss 3T3 fibroblasts to stimulate tyrosine phosphorylation of p125(FAK) and paxillin and cell growth.
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PMID:Sphingosine 1-phosphate stimulates rho-mediated tyrosine phosphorylation of focal adhesion kinase and paxillin in Swiss 3T3 fibroblasts. 918 7

Tyrosine phosphorylation of paxillin by the focal adhesion kinase (FAK) has been implicated as a signal transduction mechanism associated with cell adhesion and cytoskeletal reorganization. The potential role of serine phosphorylation of paxillin in these events has not been well characterized. In this study we have examined the phosphorylation profile of paxillin both in vitro and in vivo. By using glutathione S-transferase-paxillin fusion proteins in precipitation-kinase assays in vitro we observed that a fusion protein spanning amino acid residues 54-313 of paxillin, and containing a FAK-binding site, precipitated substantial serine kinase activity as well as FAK activity from a smooth-muscle lysate. Together these kinases phosphorylated paxillin on tyrosine residue 118, a site that has been identified previously as a target for FAK phosphorylation, and on serine residues 188 and/or 190. The binding site for the serine kinase, the identity of which is currently unknown, was further mapped to residues 168-191 of paxillin. To assess the physiological relevance of these sites phosphorylated in vitro, the profile of paxillin phosphorylation in vivo stimulated by seeding fibroblasts on fibronectin was characterized. As expected, plating cells on fibronectin enhanced the tyrosine phosphorylation of paxillin. However, 96% of the phosphorylation of paxillin occurred on serine residues. Comparison by two-dimensional phosphopeptide analyses indicated that the major sites of tyrosine and serine phosphorylation detected in the assays in vitro co-migrate with phosphopeptides derived from paxillin phosphorylated in vivo in response to plating cells on fibronectin. These findings support a role for both tyrosine and serine kinases in the signal transduction pathway linking integrin activation to paxillin phosphorylation.
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PMID:Adhesion of fibroblasts to fibronectin stimulates both serine and tyrosine phosphorylation of paxillin. 923 Jan 16

The inappropriate activation of protein-tyrosine kinases (PTKs) has been associated with initiation and progression of several types of human cancers. We therefore postulated that immortalization by DNA tumor viruses results in the induction of PTKs fundamental to these processes. An RT-PCR-based screen was thus used to identify PTKs that were abundantly expressed in HPV-18-immortalized epithelial cells and HPV-containing carcinoma cell lines. One of the genes isolated in this screen was the focal adhesion kinase (FAK; pp125FAK), a cytoplasmic protein kinase that is activated in v-src transformed cells or by stimulation with mitogenic polypeptides. FAK also becomes catalytically active upon integrin engagement with extracellular matrix proteins, such as fibronectin. We found that FAK expression and activity were significantly elevated in HPV-18 E6/E7-immortalized human genital epithelial cells relative to their primary cell counterparts. Protein expression and tyrosine phosphorylation of the putative FAK substrate, paxillin, were also notably increased upon HPV-18 immortalization of genital epithelial cells and in HPV-containing cervical carcinoma cell lines. Most significantly, these cells expressed markedly higher levels of both intracellular and extracellular fibronectin, thus providing a mechanism for activation of FAK and increased tyrosine phosphorylation of paxillin. These findings suggest a role for the integrin/FAK-mediated signaling pathway in cervical carcinogenesis and represent one of the first demonstrations of a tyrosine kinase whose activity is elevated following viral immortalization.
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PMID:Activation of the focal adhesion kinase signal transduction pathway in cervical carcinoma cell lines and human genital epithelial cells immortalized with human papillomavirus type 18. 923 61

Cell attachment to fibronectin stimulates the integrin-dependent interaction of p85-associated phosphatidylinositol (PI) 3-kinase with integrin-dependent focal adhesion kinase (FAK) as well as activation of the Ras/mitogen-activated protein (MAP) kinase pathway. However, it is not known if this PI 3-kinase-FAK interaction increases the synthesis of the 3-phosphorylated phosphoinositides (3-PPIs) or what role, if any, is played by activated PI 3-kinase in integrin signaling. We demonstrate here the integrin-dependent accumulation of the PI 3-kinase products, PI 3,4-bisphosphate [PI(3,4)P2] and PI(3,4,5)P3, as well as activation of AKT kinase, a serine/threonine kinase that can be stimulated by binding of PI(3,4)P2. The PI 3-kinase inhibitors wortmannin and LY294002 significantly decreased the integrin-induced accumulation of the 3-PPIs and activation of AKT kinase, without having significant effects on the levels of PI(4,5)P2 or tyrosine phosphorylation of paxillin. These inhibitors also reduced cell adhesion/spreading onto fibronectin but had no effect on attachment to polylysine. Interestingly, integrin-mediated Erk-2, Mek-1, and Raf-1 activation, but not Ras-GTP loading, was inhibited at least 80% by wortmannin and LY294002. In support of the pharmacologic results, fibronectin activation of Erk-2 and AKT kinases was completely inhibited by overexpression of a dominant interfering p85 subunit of PI 3-kinase. We conclude that integrin-mediated adhesion to fibronectin results in the accumulation of the PI 3-kinase products PI(3,4)P2 and PI(3,4,5)P3 as well as the PI 3-kinase-dependent activation of the kinases Raf-1, Mek-1, Erk-2, and AKT and that PI 3-kinase may function upstream of Raf-1 but downstream of Ras in integrin activation of Erk-2 MAP and AKT kinases.
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PMID:Phosphatidylinositol 3-kinase is required for integrin-stimulated AKT and Raf-1/mitogen-activated protein kinase pathway activation. 923 99


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