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)

Cell adhesion kinase-beta (CAK-beta) is a protein tyrosine kinase of the focal adhesion kinase subfamily, which contains large amino- and carboxyl-terminal domains. We studied the tissue distribution of CAK-beta and its mRNA by immunohistochemical staining and in situ hybridization. In rat brain, CAK-beta was mainly found in the medulla whereas CAK-beta mRNA was expressed in most neurons, especially pyramidal cells and Purkinje cells. In the small intestine, CAK-beta protein and mRNA were detected in the absorptive epithelial cells, and the protein was concentrated in the brush border. Double immunostaining for CAK-beta and actin showed that they co-localized in the brush border of small intestine cells. Immunoelectron micrography revealed that the anti-CAK-beta antibody localized within microvilli. In the kidney, the protein was mainly expressed in proximal tubular cells, which have well developed microvilli, although CAK-beta mRNA was observed in most urinary tubular cells. In other tissues, the ciliated cells of the epididymis strongly expressed CAK-beta mRNA and CAK-beta localized in the cilia. In addition, alpha- and beta-tubulin were identified in the rat brain lysates immunoprecipitated with anti-CAK-beta antibody. The present results demonstrate that CAK-beta is present at relatively high levels in cilia, axons, and microvilli. This suggests that CAK-beta may play important roles in the functions of these structures or that the CAK-beta-related signaling pathway is closely associated with cytoskeletal components.
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PMID:Restricted expression of cell adhesion kinase-beta in rat tissues. 900 42

Focal adhesion kinase (pp125(FAK)) is a protein tyrosine kinase that is localized to focal adhesions in many cell types and which undergoes tyrosine phosphorylation after integrin binding to extracellular matrix. In some cells the C-terminal non-catalytic domain of pp125(FAK) is expressed as a separate protein referred to as FRNK (FAK-related, non-kinase). We have previously shown that overexpression of FRNK inhibits tyrosine phosphorylation of pp125(FAK) and its substrates as well as inhibiting cell spreading on fibronectin. In this report we identify Ser148 and Ser151 as residues in FRNK that are phosphorylated after tyrosine phosphorylation of pp125(FAK) and in response to integrin binding to fibronectin. Tyrosine phosphorylation of pp125(FAK) appears to be an early event after integrin occupancy, and serine phosphorylation of FRNK occurs significantly later. Treatment of fibroblasts with a series of protein kinase A inhibitors delayed serine phosphorylation of FRNK as well as cell spreading on fibronectin and tyrosine phosphorylation of pp125(FAK). However, these PKA inhibitors are unlikely to delay cell spreading simply by preventing serine phosphorylation of FRNK, as overexpression of FRNK containing mutations of Ser148 and Ser151 either singly or jointly to either alanine or glutamate residues did not significantly alter the ability of FRNK to act as an inhibitor of pp125(FAK).
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PMID:Identification of integrin-stimulated sites of serine phosphorylation in FRNK, the separately expressed C-terminal domain of focal adhesion kinase: a potential role for protein kinase A. 916 50

Focal adhesion kinase (FAK) has been overexpressed in insect cells using a baculovirus expression system. A recombinant baculovirus was generated which contains the mouse FAK cDNA cloned into a histidine tag transfer vector. Synthesis of the immunoreactive recombinant protein (baculovirus focal adhesion kinase (BFAK) Mr approximately 125,000) in infected Sf9 cells was detected 23 h postinfection, with maximal accumulation occurring at 48 h postinfection. BFAK constituted 5.4% of total soluble protein in the insect cell lysate and represented 19 mg/liter culture (approximately 2 x 10(9) cells). The enzyme was active as a protein tyrosine kinase in both SF9 cells and in vitro. Purification to near homogeneity was achieved by nickel chelation chromatography. A yield of 5 mg of purified active BFAK was consistently produced from 1 liter of infected insect cells. BFAK tyrosine kinase activity was characterized physically using poly(Glu-Tyr) as a substrate. BFAK activity required the presence of a divalent cation and exhibited a preference for Mn2+ over Mg2+. Maximal tyrosine kinase activity was attained at pH 7.2. Steady-state kinetic analysis with respect to ATP concentration did not conform to simple Michaelis-Menten kinetics and exhibited a Hill coefficient of much less than 1. Km values for ATP using native and autophosphorylated BFAK were 6.7 +/- 1.0 and 4.3 +/- 0.2 microM, respectively. Kcat values were 13.9 +/- 1.9 and 8.9 +/- 0.3 nmol/min/mg BFAK. Steady-state kinetics with respect to the peptide substrate did fit the Michaelis-Menten equation and exhibited a Km value of 2.4 +/- 0.3 micro/ml.
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PMID:Expression, purification and characterization of focal adhesion kinase using a baculovirus system. 917 76

JAK3 is a protein tyrosine kinase that specifically associates with the common gamma chain (gammac), a shared subunit of receptors for interleukin (IL) 2, 4, 7, 9, and 15. Patients deficient in either JAK3 or gammac presented with virtually identical forms of severe combined immunodeficiency (SCID), underscoring the importance of the JAK3-gammac interaction. Despite the key roles of JAK3 and gammac in lymphocytic development and function, the molecular basis of this interaction remains poorly understood. In this study, we have characterized the regions of JAK3 involved in gammac association. By developing a number of chimeric JAK3-JAK2 constructs, we show that the binding specificity to gammac can be conferred to JAK2 by transferring the N-terminal domains of JAK3. Moreover, those JAK3-JAK2 chimeras capable of binding gammac were also capable of reconstituting IL-2 signaling as measured by inducible phosphorylation of the chimeric JAK3-JAK2 protein, JAK1, the IL-2 receptor beta chain, and signal transducer and activator of transcription 5A. Subsequent deletion analyses of JAK3 have identified the N-terminal JH7-6 domains as a minimal region sufficient for gammac association. Furthermore, expression of the mutant containing only the JH7-6 domains effectively competed with full-length JAK3 for binding to gammac. We conclude that the JH7-6 domains of JAK3 are necessary and sufficient for gammac association. These studies offer clues toward a broader understanding of JAK-mediated cytokine signaling and may provide a target for the development of novel therapeutic modalities in immunologically mediated diseases.
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PMID:The amino terminus of JAK3 is necessary and sufficient for binding to the common gamma chain and confers the ability to transmit interleukin 2-mediated signals. 919 65

Cell adhesion kinase beta (CAKbeta) is a protein tyrosine kinase closely related to focal adhesion kinase (FAK) in structure. CAKbeta contains two proline-rich sequences within its C-terminal region. Since proline-rich sequences present in the corresponding region of FAK are known to mediate protein-protein interactions by binding to SH3 domains, we investigated binding of CAKbeta to a panel of SH3 domains. Affinity precipitation from rat brain lysate revealed selective interactions of CAKbeta with glutathione S-transferase (GST)-fused SH3 domains of p130(Cas)(Cas)-related proteins and Graf. Mutational analysis indicated that the proline-rich sequences of CAKbeta mediate this interaction. Each of the two proline-rich sequences fused to GST bound directly to these SH3 domains in dot blot analysis. A competitive binding assay revealed that the first proline-rich sequence of CAKbeta preferentially associated with the SH3 domain of Cas. The second proline-rich sequence of CAKbeta bound to the SH3 domain of Graf with higher specificity than the corresponding proline-rich sequence of FAK. Finally, we showed co-immunoprecipitation of CAKbeta with Graf from rat brain lysate. These results indicate that CAKbeta associates in vivo with Graf through its SH3 domain.
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PMID:Interaction of two proline-rich sequences of cell adhesion kinase beta with SH3 domains of p130Cas-related proteins and a GTPase-activating protein, Graf. 949 93

Cell adhesion to the extracellular matrix (ECM) has been implicated in apoptosis in anchorage-dependent cell types. We recently found that a peptide derived from fibronectin (termed III14-2) inhibits the integrin-mediated cell adhesion to ECM. Using this antiadhesive peptide and a variety of ECM proteins, we show here a critical role of the integrin-ECM protein interaction in apoptotic regulation of human umbilical vein endothelial cells (HUVEC). HUVEC in suspension underwent apoptosis under the serum-free conditions, as judged by nuclear and DNA fragmentations. This apoptosis was suppressed to varying degrees when alpha 5 beta 1, alpha v beta 3, and alpha 2 beta 1 integrins were occupied with either soluble or immobilized ECM proteins such as fibronectin, vitronectin, and type I collagen, respectively. Peptide III14-2, which had no effect by itself on the HUVEC apoptosis, disrupted the ligation of alpha 5 beta 1 and alpha v beta 3 but no alpha 2 beta 1 and ultimately led the cells to apoptosis, indicating that this antiadhesive peptide indirectly induces apoptosis by blocking cell survival signal delivered from alpha 5 beta 1 and alpha v beta 3 integrins. Genistein, a protein tyrosine kinase inhibitor, slightly reduced the rescuing effect of fibronectin, whereas sodium orthovanadate and bombesin, which increase in the level of protein tyrosine phosphorylation, made HUVEC less susceptible to apoptosis and blocked the effect of peptide III14-2. HUVEC adhesion to fibronectin substrate raised the tyrosine phosphorylation level of focal adhesion kinase and the expression of cytoprotective Bcl-2 protein, both of which were reversed by the antiadhesive effect of peptide III14-2. Thus, the opposing effects of ECM proteins, including fibronectin and vitronectin, and peptide III14-2 on HUVEC apoptosis appear to be due to the opposing effects of these factors on the signaling pathway which includes tyrosine phosphorylation of FAK and Bcl-2 expression.
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PMID:Modulation of apoptotic cell death by extracellular matrix proteins and a fibronectin-derived antiadhesive peptide. 966 6

We have previously shown that integrin-dependent tyrosine phosphorylation of p130Cas (Cas) could be induced in a mouse fibroblast cell line that does not express focal adhesion kinase p125FAK (FAK). By analyzing FAK-deficient (FAK-/-) cells transiently expressing Cas mutant proteins, we demonstrate here that the Src homology 3 (SH3) domain of Cas is indispensable for adhesion-mediated Cas phosphorylation in this mutant cell line. While the FAK directly binds to Cas-SH3, our findings imply that SH3-binding molecule(s) other than FAK might regulate Cas phosphorylation, at least in FAK-/- cells. In this regard, we observed that FAK-/- cells expressed cell adhesion kinase beta (CAKbeta), a protein tyrosine kinase of the FAK subfamily. CAKbeta expressed by FAK-/- cells was associated in vivo with Cas in a Cas-SH3-dependent manner. Moreover, integrin stimulation induces tyrosine phosphorylation of CAKbeta in FAK-/- cells. Thus, our results suggest that CAKbeta contributes to integrin-mediated signal transduction in place of FAK in FAK-deficient cells.
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PMID:Integrin-mediated signal transduction in cells lacking focal adhesion kinase p125FAK. 972 Sep 24

Abnormal beta1 integrin receptor function may contribute to the continuous proliferation and abnormal circulation of malignant hematopoietic progenitors in chronic myelogenous leukemia (CML). Previous studies suggest that abnormal integrin function in CML progenitors is related to the presence of the BCR/ABL oncogene. BCR/ABL may alter integrin function in CML by phosphorylating cytoskeletal and/or signaling proteins important for normal integrin function. We evaluated the effect of Tyrphostin AG957, a protein tyrosine kinase (PTK) inhibitor which has activity against the p210BCR/ABL kinase, on beta1 integrin function in CML progenitors. Incubation of CML marrow CD34+HLA-DR+ cells with Tyrphostin AG957 at concentrations that did not affect colony-forming cells (CFC) viability, but which partly inhibited p210BCR/ABL kinase activity, significantly increased CML CFC adhesion to stroma and alpha4beta1 and alpha5beta1 integrin binding fragments of fibronectin (FN). CML CFC proliferation, unlike that of normal CFC, is not inhibited following integrin receptor engagement with FN or anti-integrin antibodies. AG957 did not alter CML CFC proliferation by itself, but resulted in significant inhibition of CML CFC proliferation following integrin engagement. Another PTK inhibitor, Tyrphostin AG555, which does not have anti-p210BCR/ABL kinase activity, did not affect CML CFC adhesion or proliferation. Neither AG957 nor AG555 affected normal CFC adhesion or proliferation. In BCR/ABL expressing cells, AG957 partially inhibited phosphorylation of several proteins that are BCR/ABL PTK substrates and are involved in normal integrin signaling. These studies suggest that abnormal tyrosine phosphorylation may play an important role in defective integrin function in CML progenitors.
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PMID:Tyrphostin AG957, a tyrosine kinase inhibitor with anti-BCR/ABL tyrosine kinase activity restores beta1 integrin-mediated adhesion and inhibitory signaling in chronic myelogenous leukemia hematopoietic progenitors. 982 45

Fc gamma receptors on monocytes/macrophages play an important role in both host defense and autoimmune disorders. Fc gamma receptor signaling can lead to such downstream events as phagocytosis and the release of intracellular cytokines and reactive oxygen species. Freshly isolated human monocytes express two major classes of Fc gamma receptor proteins, Fc gamma RI (CD64) and Fc gamma RII (CD32). Crosslinking of Fc gamma RI and Fc gamma RII gives rise to rapid and transient phosphorylation of multiple monocyte intracellular proteins including proteins of 40, 68-72, 75-85, 95, and 115-165 kDa. A 72-kDa protein was earlier identified as the tyrosine kinase Syk. Here we identify one of the proteins in the 115- to 165-kDa cluster as FAK, a protein tyrosine kinase localized to focal adhesions. A 68-kDa phosphoprotein was identified as paxillin, a cytoskeleton associated substrate for tyrosine kinases, and a 95-kDa protein was found to be the proto-oncogene product Vav. The Src family protein tyrosine kinase Fgr (p58) also displayed enhanced tyrosine phosphorylation after Fc gamma RI and Fc gamma RII crosslinking. Although Fc gamma RIIA utilizes tyrosines within its own cytoplasmic domain for signaling while Fc gamma RI utilizes the cytoplasmic tyrosines of its associated gamma subunit, our results indicate sharing of several proteins for signaling in monocytes by these Fc receptors. These molecules include three distinct classes of tyrosine kinases, Syk, FAK, and Fgr, and the functionally diverse proteins Vav and paxillin.
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PMID:Activation of three classes of nonreceptor tyrosine kinases following Fc gamma receptor crosslinking in human monocytes. 988 53

Focal adhesion kinase (pp125FAK or FAK) is a protein tyrosine kinase which is associated with intracellular signalling cascades which are initiated when the integrin family of cell adhesion molecules engage extracellular matrix molecules. In cultured cells, this molecule is physically associated with focal adhesions, which are well-defined regions of intimate cell-to-substratum adhesion. In this location, it interacts with other proteins of the focal adhesion to activate intracellular signalling events associated with cell adhesion. The in vitro expression of FAK and its level of phosphorylation appear to be related to several physiological phenomena, including cell spreading, cell differentiation, cell locomotion and cell death. Because these phenomena are all of critical importance during morphogenesis, and because FAK is expressed in embryonic cells, evidence has been accumulating to indicate that FAK may be an important modulator of developmental processes. In this review, this evidence is surveyed together with evidence from analogous situations, such as tumour cell migration and invasiveness. Although evidence suggesting a role for FAK in morphogenesis is accumulating, current uncertainties regarding its cytoplasmic location and its molecular interactions in vivo make it difficult to reach definitive conclusions regarding the significance of its contributions to developmental processes.
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PMID:Potential roles for focal adhesion kinase in development. 992 29


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