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)

Integrin alpha 3A cytoplasmic tail phosphorylation was mapped to amino acid S1042, as determined by mass spectrometry, and confirmed by mutagenesis. This residue occurs within a "QPSXXE" motif conserved in multiple alpha chains (alpha 3A, alpha 6A, alpha 7A), from multiple species. Phosphorylation of alpha 3A and alpha 6A did not appear to be directly mediated by protein kinase C (PKC) alpha, beta, gamma, delta, epsilon, zeta, or mu, or by any of several other known serine kinases, although PKC has an indirect role in promoting phosphorylation. A S1042A mutation did not affect alpha 3-Chinese hamster ovary (CHO) cell adhesion to laminin-5, but did alter 1) alpha 3-dependent tyrosine phosphorylation of focal adhesion kinase and paxillin (in the presence or absence of phorbol 12-myristate 13 acetate stimulation), and p130(CAS) (in the absence of phorbol 12-myristate 13 acetate stimulation), 2) the shape of cells spread on laminin-5, and 3) alpha 3-dependent random CHO cell migration on laminin-5. In addition, S1042A mutation altered the PKC-dependent, ligand-dependent subcellular distribution of alpha 3 and F-actin in CHO cells. Together, the results demonstrate clearly that alpha 3A phosphorylation is functionally relevant. In addition, the results strongly suggest that alpha 3 phosphorylation may regulate alpha 3 integrin interaction with the cytoskeleton.
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PMID:Phosphorylation of a conserved integrin alpha 3 QPSXXE motif regulates signaling, motility, and cytoskeletal engagement. 1117 20

Desensitization and phosphorylation of the endogenous angiotensin II AT(1) receptor were studied in clone 9 liver cells. Agonist activation of AT(1) receptors blunted the response to subsequent addition of angiotensin II. Partial inhibition of the angiotensin II-induced calcium response was observed when cells were pretreated with dibutyryl cyclic AMP, tetradecanoyl phorbol acetate (TPA), vasopressin, or lysophosphatidic acid. All of these desensitization processes were associated with receptor phosphorylation. Angiotensin II-induced AT(1) receptor phosphorylation was partially blocked by the protein kinase C inhibitor bisindolylmaleimide I and by phosphoinositide 3-kinase inhibitors (wortmannin and LY294002); the actions of these inhibitors were not additive. Pertussis toxin pretreatment of cells also partially inhibited angiotensin II-induced AT(1) receptor phosphorylation. TPA-induced AT(1) receptor phosphorylation was completely blocked by bisindolylmaleimide I. AT(1) receptor phosphorylation was also induced by vasopressin and lysophosphatidic acid, and these effects were partially inhibited by bisindolylmaleimide I. Angiotensin II increased Akt/PKB (protein kinase B) phosphorylation and protein kinase C membrane association. The effect on Akt/PKB phosphorylation was blocked by phosphoinositide 3-kinase inhibitors. These findings indicate that clone 9 cells exhibit both homologous and heterologous desensitization in association with AT(1) receptor phosphorylation. In these hepatic cells, angiotensin II-induced receptor phosphorylation involves pertussis toxin-sensitive and -insensitive G proteins, and is mediated in part through protein kinase C and phosphoinositide 3-kinase.
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PMID:Angiotensin AT(1) receptor phosphorylation and desensitization in a hepatic cell line. Roles of protein kinase c and phosphoinositide 3-kinase. 1117 53

An ester hydrolase (ABL) has been isolated from a strain of Arthrobacter species (RRLJ-1/95) maintained in the culture collection of this laboratory. The purified enzyme has a specific activity of 1700 U/mg protein and is found to be composed of a single subunit (Mr 32,000), exhibiting both lipase and esterase activities shown by hydrolysis of triglycerides and p-nitrophenyl acetate respectively. Potential application of the enzyme concerns the asymmetrisation of prochiral 2-benzyl-1,3-propanediol esters besides enantioselective hydrolysis of alkyl esters of unsubstituted and substituted 1-phenyl ethanols.
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PMID:Purification and characterisation of an ester hydrolase from a strain of Arthrobacter species: its application in asymmetrisation of 2-benzyl-1,3-propanediol acylates. 1124 19

Treatment of cultured human hepatoma HepG2 cells with the protein kinase C (PKC) activator, 12-O-tetradecanoylphorbol-13-acetate (TPA), results in an increase in tyrosine phosphorylation of several proteins, including the focal adhesion kinase (FAK) and paxillin using anti-phosphotyrosine Western blotting and immunoprecipitation. However, when cells are in suspension or in the presence of cytochalasin D which disrupts the intracellular network of actin microfilaments, TPA loses its ability to stimulate tyrosine phosphorylation of FAK and paxillin but it still activates mitogen-activated protein kinase (MAPK) and induces PKC translocation from cytosol to the membrane in HepG2 cells. On the other hand, PD98059, a specific inhibitor of mitogen-activated protein kinase kinase, blocks TPA-induced MAPK activation but has no effect on TPA-induced tyrosine phosphorylation. Our findings suggest that TPA-induced tyrosine phosphorylation of FAK and paxillin in human hepatoma cells is PKC dependent and requires the integrity of the cell cytoskeleton but is uncoupled to the signal transduction pathway of PKC leading to the translocation of PKC and MAPK activation.
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PMID:Protein kinase C-mediated tyrosine phosphorylation of paxillin and focal adhesion kinase requires cytoskeletal integrity and is uncoupled to mitogen-activated protein kinase activation in human hepatoma cells. 1128 49

Growth hormone (GH) initiates its cellular action by properly dimerizing GH receptor (GHR). A substantial fraction of circulating GH is complexed with a high-affinity GH-binding protein (GHBP) that in many species can be generated by GHR proteolysis and shedding of the receptor's ligand-binding extracellular domain. We previously showed that this proteolysis 1) can be acutely promoted by the phorbol ester phorbol 12-myristate 13-acetate (PMA), 2) requires a metalloprotease activity, 3) generates both shed GHBP and a membrane-associated GHR transmembrane/cytoplasmic domain remnant, and 4) results in down-regulation of GHR abundance and GH signaling. Using cell culture model systems, we now explore the effects of GH treatment on inducible GHR proteolysis and GHBP shedding. In human IM-9 lymphocytes, which endogenously express GHRs, and in Chinese hamster ovary cells heterologously expressing wild-type or cytoplasmic domain internal deletion mutant rabbit GHRs, brief exposure to GH inhibited PMA-induced GHR proteolysis (receptor loss and remnant accumulation) by 60-93%. PMA-induced shedding of GHBP from Chinese hamster ovary transfectants was also inhibited by 70% in the presence of GH. The capacity of GH to inhibit inducible GHR cleavage did not rely on JAK2-dependent GH signaling, as evidenced by its continued protection in JAK2-deficient gamma2A rabbit GHR cells. The GH concentration dependence for inhibition of PMA-induced GHR proteolysis paralleled that for its promotion of receptor dimerization (as monitored by formation of GHR disulfide linkage). Unlike GH, the GH antagonist, G120K, which binds to but fails to properly dimerize GHRs, alone did not protect against PMA-induced GHR proteolysis; G120K did, however, antagonize the protective effect of GH. Our data suggest that GH inhibits PMA-induced GHR proteolysis and GHBP shedding by inducing GHR dimerization and that this effect does not appear to be related to GH site 1 binding, GHR internalization, or GHR signaling. The implications of these findings with regard to GH signaling and GHR down-regulation are discussed.
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PMID:Growth hormone (GH)-induced dimerization inhibits phorbol ester-stimulated GH receptor proteolysis. 1130 89

Carbachol treatment resulted in increased phosphorylation on tyrosine of PKCdelta immunoprecipitated from rat pancreatic acinar cells. The Ca2+-dependent tyrosine kinase PYK2 coimmunoprecipitated with PKCdelta from carbachol-exposed cells and also exhibited increased tyrosine phosphorylation. Tyrosine phosphorylation of both PKCdelta and PYK2 was concentration-dependent with respect to carbachol, and rapid, reaching maximal levels by 5 min of treatment. Exposure of acinar cells to phorbol myristate acetate (PMA), a phorbol ester activator of PKCdelta, also resulted in increased phosphorylation of PKCdelta and PYK2 isolated using anti-PKCdelta immunoprecipitation. These results are suggestive of a physical and functional interaction between PKCdelta and PYK2 following muscarinic stimulation in the pancreatic acinar cell.
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PMID:Carbachol stimulates TYR phosphorylation and association of PKCdelta and PYK2 in pancreas. 1135 32

A peer-verified, solid-phase extraction (SPE)/anion exchange liquid chromatographic method is presented for the determination of niacin in milk-based and soy-based infant formula. Analysis is in 3 steps: test sample digestion, extraction/cleanup, and liquid chromatography (LC). Digestion uses a standard AOAC digestion procedure that involves autoclaving at 121 degrees C for 45 min in (1 + 1) H2SO4 to free endogenous niacin from protein and to convert added niacinamide to niacin. The digest solution is adjusted to pH 6.5 with 7.5M NaOH. Acidification to pH <1.0 with (1 + 1) H2SO4 precipitates the protein. The clarified solution is then filtered, and the filtrate is brought to volume. SPE of niacin is accomplished by passing an aliquot of the digest solution through an aromatic sulfonic acid-SPE (ArSCX-SPE) column. After the column is washed with methanol and water to remove extraneous material, the niacin is eluted with 0.25M sodium acetate/acetic acid buffer at pH 5.6. An anion-exchange polystyrene-divinylbenzene column with 0.1 M sodium acetate/acetic acid buffer at pH 4.0 is used for LC. Niacin is determined by UV detection at 260 nm. A standard curve is prepared by passing known amounts of niacin through the ArSCX-SPE columns used for niacin extraction. The following values for x and relative standard deviation (RSD) were obtained for National Institute of Standards and Technology Standard Reference Material (NIST SRM) 1846 Infant Formula with a certified value for niacin of 63.3 +/- 7.6 microg/g: Submitting laboratory.-- x = 59.7 +/- 4.0 microg/g; RSD = >6.7%; confidence interval (CI) = +/- 1.4 microg/g; n = 27. Peer laboratory.--x = 56.6 +/- 6.6 microg/g; RSD = >11.7%; CI =+/- 4.1 microg/g; n = 8.
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PMID:Determination of niacin in infant formula by solid-phase extraction and anion-exchange liquid chromatography. 1141 44

To examine the effect of exercise and adrenergic blockade on lymphocyte cytokine production, six men ingested either a placebo (control) or an alpha- (prazosin hydrochloride) and beta-adrenoceptor antagonist (timolol malate) capsule (blockade, or BLK) 2 h before performing 19 +/- 1 min of supine bicycle exercise at 78 +/- 3% peak pulmonary uptake. Blood was collected before and after exercise, stimulated with phorbol 12-myristate 13-acetate and ionomycin, and surface stained for T (CD3(+)) and natural killer [NK (CD3(-)CD56(+))] lymphocyte surface antigens. Cells were permeabilized, stained for the intracellular cytokines interleukin (IL)-2 and interferon (IFN)-gamma, and analyzed using flow cytometry. BLK had no effect on the resting concentration of stimulated cytokine-positive T and NK lymphocytes or the amount of cytokine they were producing. Exercise resulted in an increase (P < 0.05) in the concentration of stimulated T and NK lymphocytes producing cytokines in the circulation, but these cells produced less (P < 0.05) cytokine post- compared with preexercise. BLK attenuated (P < 0.05) the elevation in the concentration of lymphocytes producing cytokines during exercise; however, BLK did not affect the amount of IL-2 and IFN-gamma produced. These results suggest that adrenergic stimulation contributes to the exercise-induced increase in the concentration of lymphocytes in the circulation; however, it does not appear to be responsible for the exercise-induced suppression in cytokine production.
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PMID:Effect of adrenergic blockade on lymphocyte cytokine production at rest and during exercise. 1154 60

The signal transduction pathway linking physiological concentrations of [Arg(8)]vasopressin (AVP) to an increase in frequency of Ca(2+) spiking was examined in confluent cultures of A7r5 vascular smooth muscle cells. Immunoprecipitation/Western blot studies revealed a robust increase in tyrosine phosphorylation of the non-receptor tyrosine kinase, PYK2, in A7r5 cells treated with 4beta-phorbol 12-myristate 13-acetate or ionomycin. 100 pm AVP also induced PYK2 tyrosine phosphorylation, and this effect was inhibited by protein kinase C inhibitors Ro-31-8220 (1-10 microm) or chelerythrine chloride (1-20 microm). In fura-2-loaded A7r5 cells, the stimulation of Ca(2+) spiking by 100 pm AVP or 1 nm 4beta-phorbol 12-myristate 13-acetate was completely blocked by PP2 (10 microm, a Src family kinase inhibitor). Salicylate (20 mm, recently identified as a PYK2 inhibitor) and the tyrosine kinase inhibitor, tyrphostin A47 (50 microm), but not its inactive analog, tyrphostin A63, also blocked AVP-stimulated Ca(2+) spiking. PYK2 phosphorylation was inhibited by both PP2 and salicylate, whereas tyrphostin A47 failed to inhibit PYK2 tyrosine phosphorylation. ERK1/2 kinases did not appear to be involved because 1) 100 pm AVP did not appreciably increase ERK1/2 phosphorylation and U-0126 (2.5 microm) did not inhibit AVP-stimulated Ca(2+) spiking; and 2) epidermal growth factor (10 nm) robustly stimulated ERK1/2 phosphorylation but did not induce Ca(2+) spiking. Delayed rectifier K(+) channels may mediate the PYK2 activity because Kv1.2 channel protein co-immunoprecipitated with PYK2 and tyrosine phosphorylation of Kv1.2 was stimulated by AVP and inhibited by Ro-31-8220, PP2, and salicylate but not tyrphostin A47. Our findings are consistent with a role for PYK2 and phosphorylation of K(+) channels in the stimulation of Ca(2+) spiking by physiological concentrations of AVP.
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PMID:Signal transduction of physiological concentrations of vasopressin in A7r5 vascular smooth muscle cells. A role for PYK2 and tyrosine phosphorylation of K+ channels in the stimulation of Ca2+ spiking. 1173 73

Interferon-gamma (IFN-gamma) induced intercellular adhesion molecule-1 (ICAM-1) expression in human NCI-H292 epithelial cells, as shown by enzyme-linked immunosorbent assay and immunofluorescence staining. The enhanced ICAM-1 expression resulted in increased adhesion of U937 cells to NCI-H292 cells. Tyrosine kinase inhibitors (genistein or herbimycin), Src family inhibitor (PP2), or a phosphatidylinositol-phospholipase C inhibitor (U73122) attenuated the IFN-gamma-induced ICAM-1 expression. Protein kinase C (PKC) inhibitors (staurosporine or Ro 31-8220) also inhibited IFN-gamma-induced response. 12-O-Tetradecanoylphorbol-13-acetate (TPA), a PKC activator, stimulated ICAM-1 expression; this effect was inhibited by tyrosine kinase or Src inhibitor. ICAM-1 promoter activity was enhanced by IFN-gamma and TPA in cells transfected with pIC339-Luc, containing the downstream NF-kappaB and gamma-activated site (GAS) sites, but not in cells transfected with GAS-deletion mutant, pIC135 (DeltaAP2). Electrophoretic gel mobility shift assay demonstrated that GAS-binding complexes in IFN-gamma-stimulated cells contained STAT1alpha. The IFN-gamma-induced ICAM-1 promoter activity was inhibited by tyrosine kinase inhibitors, a phosphatidylinositol-phospholipase C inhibitor, or PKC inhibitors, and the TPA-induced ICAM-1 promoter activity was also inhibited by tyrosine kinase inhibitors. Cotransfection with a PLC-gamma2 mutant inhibited IFN-gamma- but not TPA-induced ICAM-1 promoter activity. However, cotransfection with dominant negative mutants of PKCalpha or c-Src inhibited both IFN-gamma- and TPA-induced ICAM-1 promoter activity. The ICAM-1 promoter activity was stimulated by cotransfection with wild type PLC-gamma2, PKCalpha, c-Src, JAK1, or STAT1. An immunocomplex kinase assay showed that both IFN-gamma and TPA activated c-Src and Lyn activities and that these effects were inhibited by staurosporine and herbimycin. Thus, in NCI-H292 epithelial cells, IFN-gamma activates PLC-gamma2 via an upstream tyrosine kinase to induce activation of PKC-alpha and c-Src or Lyn, resulting in activation of STAT1alpha, and GAS in the ICAM-1 promoter, followed by initiation of ICAM-1 expression and monocyte adhesion.
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PMID:Interferon-gamma-induced epithelial ICAM-1 expression and monocyte adhesion. Involvement of protein kinase C-dependent c-Src tyrosine kinase activation pathway. 1175 11


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