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)

Previous studies have shown that collagen gel overlay induced selective proteolysis of focal adhesion complex proteins in Madin-Darby canine kidney (MDCK) cells. In this study, we examined whether morphological and biochemical changes were present in cells cultured on collagen gel. We found that focal adhesion complex proteins, including focal adhesion kinase (FAK), talin, paxillin, and p130cas, but not vinculin, were decreased within 1 h when MDCK cells were cultured on collagen gel. Collagen gel-induced selective decrease of focal adhesion proteins was observed in all lines of cells examined, including epithelial, fibroblastic, and cancer cells. Matrigel also induced selective down-regulation of focal adhesion proteins. However, cells cultured on collagen gel- or matrigel-coated dishes did not show any changes of focal adhesion proteins. These data suggest that the physical nature of the gel, i.e. the rigidity, is involved in the expression of focal adhesion proteins. The collagen gel-induced down-regulation of focal adhesion complex proteins was caused by reduction of protein synthesis and activation of proteases such as calpain. Overexpression of a dominant negative mutant of discoidin domain receptor 1 (DDR1) or FAK-related non-kinase (FRNK) did not prevent collagen gel-induced down-regulation of the focal adhesion complex protein, whereas an anti-alpha2beta1 integrin-neutralizing antibody completely blocked it. Taken together, our results indicate that the rigidity of collagen gel controls the expression of focal adhesion complex proteins, which is mediated by alpha2beta1 integrin but not DDR1.
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PMID:Rigidity of collagen fibrils controls collagen gel-induced down-regulation of focal adhesion complex proteins mediated by alpha2beta1 integrin. 1267 63

KAI1/CD82 protein is a member of the tetraspanin superfamily and has been rediscovered as a cancer metastasis suppressor. The mechanism of KAI1/CD82-mediated suppression of cancer metastasis remains to be established. In this study, we found that migration of the metastatic prostate cancer cell line Du145 was substantially inhibited when KAI1/CD82 was expressed. The expression of focal adhesion kinase (FAK) and Lyn, a Src family tyrosine kinase and substrate of FAK, was up-regulated at both RNA and protein levels upon KAI1/CD82 expression. The activation of FAK and Lyn, however, remained unchanged in Du145-KAI1/CD82 cells. As a downstream target of FAK-Lyn signaling, the p130CAS (Crk-associated substrate) protein was decreased upon the expression of KAI1/CD82. Consequently, less p130CAS-CrkII complex, which functions as a "molecular switch" in cell motility, was formed in Du145-KAI1/CD82 cells. To confirm that the p130CAS-CrkII complex is indeed important for the motility inhibition by KAI1/CD82, overexpression of p130CAS in Du145-KAI1/CD82 cells increased the formation of p130CAS-CrkII complex and largely reversed the KAI1/CD82-mediated inhibition of cell motility. Taken together, our studies indicate the following: 1) signaling of FAK-Lyn-p130CAS-CrkII pathway is altered in KAI1/CD82-expressing cells, and 2) p130CAS-CrkII coupling is required for KAI1/CD82-mediated suppression of cell motility.
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PMID:Requirement of the p130CAS-Crk coupling for metastasis suppressor KAI1/CD82-mediated inhibition of cell migration. 1273 93

Vascular endothelial growth factor (VEGF) plays a significant role in blood-brain barrier breakdown and angiogenesis after brain injury. VEGF-induced endothelial cell migration is a key step in the angiogenic response and is mediated by an accelerated rate of focal adhesion complex assembly and disassembly. In this study, we identified the signaling mechanisms by which VEGF regulates human brain microvascular endothelial cell (HBMEC) integrity and assembly of focal adhesions, complexes comprised of scaffolding and signaling proteins organized by adhesion to the extracellular matrix. We found that VEGF treatment of HBMECs plated on laminin or fibronectin stimulated cytoskeletal organization and increased focal adhesion sites. Pretreating cells with VEGF antibodies or with the specific inhibitor SU-1498, which inhibits Flk-1/KDR receptor phosphorylation, blocked the ability of VEGF to stimulate focal adhesion assembly. VEGF induced the coupling of focal adhesion kinase (FAK) to integrin alphavbeta5 and tyrosine phosphorylation of the cytoskeletal components paxillin and p130cas. Additionally, FAK and related adhesion focal tyrosine kinase (RAFTK)/Pyk2 kinases were tyrosine-phosphorylated by VEGF and found to be important for focal adhesion sites. Overexpression of wild type RAFTK/Pyk2 increased cell spreading and the migration of HBMECs, whereas overexpression of catalytically inactive mutant RAFTK/Pyk2 markedly suppressed HBMEC spreading ( approximately 70%), adhesion ( approximately 82%), and migration ( approximately 65%). Furthermore, blocking of FAK by the dominant-interfering mutant FRNK (FAK-related non-kinase) significantly inhibited HBMEC spreading and migration and also disrupted focal adhesions. Thus, these studies define a mechanism for the regulatory role of VEGF in focal adhesion complex assembly in HBMECs via activation of FAK and RAFTK/Pyk2.
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PMID:Vascular endothelial growth factor regulates focal adhesion assembly in human brain microvascular endothelial cells through activation of the focal adhesion kinase and related adhesion focal tyrosine kinase. 1284 92

Mitogen-induced changes in the actin cytoskeleton are accompanied by changes in the tyrosine phosphorylation of several proteins in focal adhesions. In this study, we have investigated the role of RAFTK (also termed Pyk2/CAK-beta), a cytoplasmic tyrosine kinase related to focal adhesion kinase (FAK), in heregulin-mediated signal transduction in breast cancer cells. Stimulation of T47D cells with heregulin (HRG) induced the tyrosine phosphorylation of RAFTK and the formation of a multiprotein complex. Maximal phosphorylation of the proteins participating in this complex occurred within 2 h of HRG stimulation. Analyses of the members of the HRG-stimulated complex revealed that RAFTK associated with p190 RhoGAP (p190), RasGAP, c-Abl as well as with the focal adhesion molecules p130cas and paxillin. c-Abl was found to be associated with RAFTK through the region of RAFTK containing amino acids 419-1009. Site-directed mutagenesis of Y881 aa within the RAFTK sequence abolished the binding of RAFTK to c-Abl, indicating that the tyrosine residue 881 of RAFTK is the c-Abl binding site within the RAFTK molecule. Overexpression of wild-type RAFTK significantly enhanced breast cancer cell invasion, while overexpression of the mutants Tyr402 or Tyr881 of RAFTK inhibited this migration. Therefore, RAFTK serves as a mediator and an integration point between focal adhesion molecules in HRG-mediated signaling in T47D breast cancer cells.
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PMID:Coupling of RAFTK/Pyk2 kinase with c-Abl and their role in the migration of breast cancer cells. 1465 52

Cell migration is a complex, highly regulated process that involves the continuous formation and disassembly of adhesions (adhesion turnover). Adhesion formation takes place at the leading edge of protrusions, whereas disassembly occurs both at the cell rear and at the base of protrusions. Despite the importance of these processes in migration, the mechanisms that regulate adhesion formation and disassembly remain largely unknown. Here we develop quantitative assays to measure the rate of incorporation of molecules into adhesions and the departure of these proteins from adhesions. Using these assays, we show that kinases and adaptor molecules, including focal adhesion kinase (FAK), Src, p130CAS, paxillin, extracellular signal-regulated kinase (ERK) and myosin light-chain kinase (MLCK) are critical for adhesion turnover at the cell front, a process central to migration.
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PMID:FAK-Src signalling through paxillin, ERK and MLCK regulates adhesion disassembly. 1474 21

The streptococcal collagen-like proteins Scl1 and Scl2 are prokaryotic members of a large protein family with domains containing the repeating amino acid sequence (Gly-Xaa-Yaa)(n) that form a collagen-like triple-helical structure. Here, we test the hypothesis that Scl variant might interact with mammalian collagen-binding integrins. We show that the recombinant Scl protein p176 promotes adhesion and spreading of human lung fibroblast cells through an alpha2beta1 integrin-mediated interaction as shown in cell adhesion inhibition assays using anti-alpha2beta1 and anti-beta1 integrins monoclonal antibodies. Accordingly, C2C12 cells stably expressing alpha2beta1 integrin as the only collagen-binding integrin show productive cell adhesion activities on p176 that can be blocked by an anti-alpha2beta1 integrin antibody. In addition, p176 promotes tyrosine phosphorylation of p125(FAK) of C2C12 cells expressing alpha2beta1 integrin, whereas parental cells do not. Furthermore, C2C12 adhesion of human lung fibroblast cells to p176 induces phosphorylation of p125FAK, p130CAS, and p68Paxillin proteins. In a domain swapping experiment, we show that integrin binds to the collagenous domain of the Scl protein. Moreover, the recombinant inserted domain of the alpha2 integrin interacts with p176 with a relatively high affinity (K(D) = 17 nm). Attempts to identify the integrin sites in p176 suggest that more than one site may be involved. These studies, for the first time, suggest that the collagen-like proteins of prokaryotes retained not only structural but also functional characteristics of their eukaryotic counterparts.
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PMID:A streptococcal collagen-like protein interacts with the alpha2beta1 integrin and induces intracellular signaling. 1564 74

Induction of humoral anti-human high molecular weight melanoma-associated antigen (anti-HMW-MAA) immunity following active specific immunotherapy is associated with a statistically significant prolongation of survival in patients with melanoma. This association does not appear to be mediated by immunological mechanisms because anti-HMW-MAA antibodies are poor mediators of complement- and cell-mediated cytotoxicity of melanoma cells. Therefore, we have been investigating nonimmunological mechanisms by which anti-HMW-MAA antibodies (Abs) affect the biology of melanoma cells. We have demonstrated that anti-HMW-MAA mAbs interfere with the interaction of HMW-MAA with extracellular matrix (ECM) components, a process known to be crucial in the early phase of melanoma metastasis. Furthermore, anti-HMW-MAA mAbs appear to block the series of signal transduction events triggered by the interaction of HMW-MAA with ECM. They include the activation of the family of Rho GTPases, p130cas, and focal adhesion kinase (FAK). These findings parallel the inhibition of the rat homologue of HMW-MAA NG2 function by anti-NG2 antibodies. Taken together, all these results provide a mechanistic explanation not only for the therapeutic effect of anti-HMW-MAA antibodies in the treatment of melanoma, but also for the function of HMW-MAA in the biology of melanoma cells. This information is expected to serve as a useful background to design effective HMW-MAA-targeted immunotherapy in patients with melanoma.
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PMID:Immunotherapy of melanoma targeting human high molecular weight melanoma-associated antigen: potential role of nonimmunological mechanisms. 1565 Feb 59

The Crk-associated tyrosine kinase substrate p130cas (CAS) is a docking protein containing an SH3 domain near its N terminus, followed by a short proline-rich segment, a large central substrate domain composed of 15 repeats of the four amino acid sequence YxxP, a serine-rich region and a carboxy-terminal domain, which possesses consensus binding sites for the SH2 and SH3 domains of Src (YDYV and RPLPSPP, respectively). The SH3 domain of CAS mediates its interaction with several proteins involved in signaling pathways such as focal adhesion kinase (FAK), tyrosine phosphatases PTP1B and PTP-PEST, and the guanine nucleotide exchange factor C3G. As a homolog of the corresponding Src docking domain, the CAS SH3 domain binds to proline-rich sequences (PxxP) of its interacting partners that can adopt a polyproline type II helix. We have determined a high-resolution X-ray structure of the recombinant human CAS SH3 domain. The domain, residues 1-69, crystallized in two related space groups, P2(1) and C222(1), that provided diffraction data to 1.1 A and 2.1 A, respectively. The crystal structure shows, in addition to the conserved SH3 domain architecture, the way in which the CAS characteristic amino acids form an atypically charged ligand-binding surface. This arrangement provides a rationale for the unusual ligand recognition motif exhibited by the CAS SH3 domain. The structure enables modelling of the docking interactions to its ligands, for example from focal adhesion kinase, and supports structure-based drug design of inhibitors of the CAS-FAK interaction.
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PMID:The 1.1 A resolution crystal structure of the p130cas SH3 domain and ramifications for ligand selectivity. 1578 59

We have previously observed that collagen IV regulates Caco-2 intestinal epithelial cell spreading and migration via Src kinase and stimulates Src-dependent tyrosine phosphorylation of p130cas. We observed that collagen IV also stimulated Src-dependent phosphorylation of both paxillin Tyr31 and paxillin Tyr118. Caco-2 transfection with paxillin or p130cas siRNAs inhibited expression of these proteins by more than 90% for at least 5 days after transfection. Although p130cas siRNA inhibited cell spreading on collagen IV by 33%, three different paxillin siRNAs did not inhibit cell spreading. p130cas siRNA did not affect Src Tyr416 or Src Tyr527 phosphorylation, FAK Tyr397 phosphorylation, or Src-dependent phosphorylation of FAK Tyr925, suggesting that p130cas did not inhibit cell spreading by altering FAK or Src activity. Rat p130cas expression after siRNA knock-out of endogenous human p130cas in Caco-2 cells reduced cell spreading inhibition by 71%. In contrast, expression of rat p130cas from which the Src-phosphorylated substrate domain was deleted did not rescue siRNA inhibition of cell spreading. Combined treatment with siRNAs to Crk and CrkL, which bind to the p130cas substrate domain, inhibited cell spreading by 54%. Both p130cas siRNA and the combined Crk/CrkL siRNAs strongly inhibited (52 and 46% inhibition, respectively) Caco-2 sheet migration on collagen IV and noticeably inhibited lamellipodial extension, whereas paxillin siRNA only inhibited migration by 18% and did not noticeably affect lamellipodial extension. These results suggest that Src may regulate Caco-2 migration on collagen IV via both p130cas and paxillin but that Src phosphorylation of p130cas is more important for this process.
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PMID:p130cas but not paxillin is essential for Caco-2 intestinal epithelial cell spreading and migration on collagen IV. 1581 76

The highly invasive behavior of glioblastoma cells contributes to the morbidity and mortality associated with these tumors. The integrin-mediated adhesion and migration of glioblastoma cells on brain matrix proteins is enhanced by stimulation with growth factors, including platelet-derived growth factor (PDGF). As focal adhesion kinase (FAK), a nonreceptor cytoplasmic tyrosine kinase, has been shown to promote cell migration in various other cell types, we analysed its role in glioblastoma cell migration. Forced overexpression of FAK in serum-starved glioblastoma cells plated on recombinant (rec)-osteopontin resulted in a twofold enhancement of basal migration and a ninefold enhancement of PDGF-BB-stimulated migration. Both expression of mutant FAK(397F) and the downregulation of FAK with small interfering (si) RNA inhibited basal and PDGF-stimulated migration. FAK overexpression and PDGF stimulation was found to increase the phosphorylation of the Crk-associated substrate (CAS) family member human enhancer of filamentation 1 (HEF1), but not p130CAS or Src-interacting protein (Sin)/Efs, although the levels of expression of these proteins was similar. Moreover downregulation of HEF1 with siRNA, but not p130CAS, inhibited basal and PDGF-stimulated migration. The phosphorylated HEF1 colocalized with vinculin and was associated almost exclusively with 0.1% Triton X-100 insoluble material, consistent with its signaling at focal adhesions. FAK overexpression promoted invasion through normal brain homogenate and siHEF1 inhibited this invasion. Results presented here suggest that HEF1 acts as a necessary and specific downstream effector of FAK in the invasive behavior of glioblastoma cells and may be an effective target for treatment of these tumors.
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PMID:HEF1 is a necessary and specific downstream effector of FAK that promotes the migration of glioblastoma cells. 1628 24


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