Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.10.2 (focal adhesion kinase)
 44,029 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Potorous tridactylis (PTK2) cells growing in culture were treated with psoralen derivatives and dividing cells were located by phase-contrast microscopy. Psoralens, light-sensitive DNA-photoadducting drugs, were reacted with mitotic chromosomes through exposure to 365-nm light from an argon laser microbeam system. It was found that following mitosis and photoreaction, cells without nuclear envelopes were produced when psoralen-treated cells received 60 light pulses over their entire chromosome complement. These 'non-nuclear membrane' cells were found to incorporate [3H]uridine and, to a lesser extent, [3H]thymidine by autoradiography. Reduction of the light exposure by half (30 near-u.v. pulses) over the entire chromosome complement in the presence of psoralen also produced non-nuclear-membrane cells as seen by light microscopy. Further examination of these cells (30 light pulses) by single-cell electron microscopy revealed that unlike the high light exposure (60 near-u.v. pulses), the low light dosage resulted in cells with membrane patches associated with their chromatin. Since neither actinomycin D nor cycloheximide impeded nuclear envelope reformation, the psoralen-DNA reaction is concluded to produce non-nuclear-membrane cells by a mechanism other than transcription or translation inhibition. The association of Golgi with areas of nuclear membrane patches gives indirect evidence of a possible Golgi contribution to the reformation of the nuclear envelope after mitosis. It is concluded that DNA plays a role in envelope reformation.
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PMID:Chromatin influence on the function and formation of the nuclear envelope shown by laser-induced psoralen photoreaction. 70 93

The potential usefulness of substrates for glycolysis and nucleic acid synthesis as tumor imaging agents was compared to that of 67Ga-citrate. In separate experiments, 3H-thymidine, 3H-uridine, 14C-2-deoxyglucose, and 67Ga-citrate were injected intraveneously into BALB/c mice with solid subcutaneous EMT-6 sarcomas. For the 3H- and 14C-labeled substrates, absolute uptakes in tumor and tumor-to-blood ratios were as high 1 hour after injection as the comparable maximum values achieved for 67Ga-citrate after 48 hours. These studies suggest that positron-labeled analogues of thymidine, uridine, and 2-deoxyglucose should be useful radiopharmaceuticals for tumor imaging by positron-emission tomography.
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PMID:Positron imaging feasibility studies: selective tumor concentration of 3H-thymidine, 3H-uridine, and 14C-2-deoxyglucose. 696 45

Leflunomide is an immunosuppressive drug capable of inhibiting T and B cell responses in vivo. A number of studies demonstrate that leflunomide functions both as a pyrimidine synthesis inhibitor and as a tyrosine kinase inhibitor. We previously reported that leflunomide inhibits LPS-stimulated B cell proliferation, cell cycle progression, and IgM secretion. This inhibition can be reversed by the addition of exogenous uridine, suggesting that leflunomide functions as a pyrimidine synthesis inhibitor in B cells. We report here that while the addition of uridine restored proliferation and IgM secretion to leflunomide-treated LPS-stimulated B cells, as determined by metabolic labeling and immunoprecipitation, it did not completely restore secretion of IgG Ab. We hypothesized that leflunomide inhibits LPS-induced IgG secretion by inhibiting tyrosine kinase activity required for isotype switch. We tested this hypothesis in a well-defined model of isotype switch, LPS plus IL-4 induction of IgG1. Leflunomide inhibited IgG1 secretion in this model in a dose-dependent manner. The signal transduction pathway utilized by IL-4 to induce IgG1 involves tyrosine phosphorylation of the IL-4 receptor, JAK1, JAK3, and STAT6 proteins induced by IL-4 binding to the IL-4R. Leflunomide diminished the tyrosine phosphorylation of JAK3 and STAT6 in the absence or presence of uridine. In gel mobility shift studies, STAT6 binding to the STAT6 DNA binding site in the IgG1 promoter decreased in the presence of leflunomide or leflunomide plus uridine. Taken together, these data suggest that leflunomide acts as a tyrosine kinase inhibitor to block IgG1 production.
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PMID:Inhibition of JAK3 and STAT6 tyrosine phosphorylation by the immunosuppressive drug leflunomide leads to a block in IgG1 production. 946 13

Uridine phosphorylase (UPase) is a key enzyme in the pyrimidine salvage pathway. It reversibly catalyzes the catabolism of uridine to uracil; controls the homeostatic regulation of uridine concentration in plasma and tissues; and plays a role in the intracellular activation of 5-fluorouracil. We cloned the murine UPase gene promoter, a 1703-bp fragment, and determined the transcription initiation sites located at +1 and +92 bp of the cDNA sequence. Through transient expression analysis of the 5'-flanking region of UPase gene, we have evaluated the promoter activity for a series of fragments with 5'- to 3'-deletion in murine breast cancer EMT-6 cells and immortalized murine fibroblast NIH 3T3 cells. Cotransfection of the UPase promoter constructs (from -1619 to -445) containing p53 binding motif with the wild-type p53 construct resulted in a significant reduction of luciferase activity; however, this effect disappeared with the additional deletion of the -445 to -274 sequence to suggest the existence in this promoter region of a putative p53 recognition element. Similar cotransfection in murine embryo fibroblasts p53-/- confirmed the inhibitory role of p53 on the UPase promoter activity. The specificity of the interaction is demonstrated by nuclear protein-specific binding to the putative p53 recognition sequence using gel mobility shift assay and DNase I footprinting analysis. These data indicate the UPase gene is a novel target of p53, and its expression is down-regulated by p53 at the promoter level.
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PMID:p53-dependent suppression of uridine phosphorylase gene expression through direct promoter interaction. 1155 67

Signaling events involving angiotensin IV (ANG IV)-mediated pulmonary artery endothelial cell (PAEC) proliferation were examined. ANG IV significantly increased upstream phosphatidylinositide (PI) 3-kinase (PI3K), PI-dependent kinase-1 (PDK-1), extracellular signal-related kinases (ERK1/2), and protein kinase B-alpha/Akt (PKB-alpha) activities, as well as downstream p70 ribosomal S6 kinase (p70S6K) activities and/or phosphorylation of these proteins. ANG IV also significantly increased 5-bromo-2'-deoxy-uridine incorporation into newly synthesized DNA in a concentration- and time-dependent manner. Pretreatment of cells with wortmannin and LY-294002, inhibitors of PI3K, or rapamycin, an inhibitor of the mammalian target of rapamycin kinase and p70S6K, diminished the ANG IV-mediated activation of PDK-1 and PKB-alpha as well as phosphorylation of p70S6K. Although an inhibitor of mitogen-activated protein kinase kinase, PD-98059, but not rapamycin, blocked ANG IV-induced phosphorylation of ERK1/2, both PD-98059 and rapamycin independently caused partial reduction in ANG IV-mediated cell proliferation. However, simultaneous treatment with PD-98059 and rapamycin resulted in total inhibition of ANG IV-induced cell proliferation. These results demonstrate that ANG IV-induced DNA synthesis is regulated in a coordinated fashion involving multiple signaling modules in PAEC.
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PMID:Activation of multiple signaling modules is critical in angiotensin IV-induced lung endothelial cell proliferation. 1222 47

Epigallocatechin-3-gallate (EGCG), a major constituent of the polyphenoids in green tea, has been reported to possess a wide range of biologic activities, including antifibrogenesis. Activated hepatic stellate cells (HSCs) are central to hepatic fibrosis, and Rho (a small GTPase)-signaling pathways have been implicated in the activation and proliferation of HSCs. In this study, we investigated the effect of EGCG on Rho-signaling pathways in activated human HSC-derived TWNT-4 cells. EGCG inhibited stress-fiber formation, an indicator of Rho activation, and changed the distribution of alpha-smooth-muscle actin. These inhibitory effects of EGCG were restored by overexpression of constitutively active Rho. A pull-down assay revealed that activated Rho (GTP-bound state) was strongly inhibited by ECGC and accompanied by suppressed phosphorylation of focal adhesion kinase, which is a regulator of Rho-signaling pathways. 5-Bromo-2'-deoxy-uridine incorporation demonstrated that ECGC (100 micromol/L suppressed cell growth by 80%, and terminal deoxynucleotidyl transferase viotin-deoxyruidine triphosphate nick end-labeling revealed that EGCG (100 micromol/L) caused apoptosis in half of the total cells. EGCG also strongly inhibited lysophoaphatidic acid (an activator of Rho) and induced phosphorylation of mitogen-activated protein kinases (Erk1/2, c-jun kinase, and p38). These findings demonstrate that EGCG regulates the structure and growth of HSCs by way of Rho-signaling pathways and suggest that EGCG has therapeutic potential in the setting of liver fibrosis.
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PMID:Epigallocatechin-3-gallate, a green-tea polyphenol, suppresses Rho signaling in TWNT-4 human hepatic stellate cells. 1597 60

Ste20-like kinase, SLK, a germinal center kinase found in kidney epithelial cells, signals to promote apoptosis. Expression of SLK mRNA and protein and kinase activity are increased during kidney development and recovery from ischemic acute renal failure. The 3'-untranslated region (3'-UTR) of SLK mRNA contains multiple adenine and uridine-rich elements, suggesting that 3'-UTR may regulate mRNA stability. This was confirmed in COS cell transient transfection studies, which showed that expression of the SLK open-reading frame plus 3'-UTR mRNA was reduced by 35% relative to the open-reading frame alone. To further characterize the SLK-3'-UTR, this nucleotide sequence was subcloned downstream of enhanced green fluorescent protein (EGFP) cDNA. In COS, 293T, and glomerular epithelial cells, expression of EGFP mRNA and protein was markedly reduced in the presence of the SLK-3'-UTR. After transfection and subsequent addition of actinomycin D, EGFP mRNA remained stable in cells for at least 6 h, whereas EGFP-SLK-3'-UTR mRNA decayed with a half-life of approximately 4 h. A region containing five AUUUA motifs within the SLK-3'-UTR destabilized EGFP mRNA. Deletion of this region from the SLK-3'-UTR, in part, restored mRNA stability. By UV cross-linking and SDS-PAGE, the SLK-3'-UTR bound to protein(s) of approximately 30 kDa in extracts of COS cells, glomerular epithelial cells, and kidney. Cotransfection of HuR (a RNA binding protein of approximately 30 kDa) increased the steady-state mRNA level of EGFP-SLK-3'-UTR but not EGFP. Thus the SLK-3'-UTR may interact with kidney RNA-binding proteins to regulate expression of SLK mRNA during kidney development and after ischemic injury.
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PMID:The 3'-untranslated region of the Ste20-like kinase SLK regulates SLK expression. 1700 24

Allogeneic immunity is one of the most potent natural immune responses. APOBEC3G (A3G) is an intracellular anti-viral factor that deaminates cytidine to uridine. Allogeneic stimulation of human CD4(+) T cells in vitro upregulated A3G mRNA and a significant correlation was found between the mixed leukocyte reaction and A3G mRNA. The mechanism of upregulation of A3G mRNA involves interaction between HLA on DC and TCR of CD4(+) T cells, which is ZAP70 and downstream ERK phosphokinase signalling dependent and induces CD40L and A3G mRNA expression in CD4(+) T cells. Alloimmune-induced A3G was found to be significantly increased in CD45RA(-), CCR5(+) and CD45RA(-)CCR7(-) subsets of effector memory T cells. In vivo studies of women alloimmunized with their partners' PBMC also showed a significant increase in A3G protein in CD4(+) T cells, CD45RO(+) memory and CCR7(-) effector memory T cells. The functional effect of allostimulation upregulating A3G mRNA was demonstrated by a significant decrease in in vitro infectivity, using GFP-labelled pseudovirus and confirmed by a decrease in HIV-1 (BaL) infection of primary CD4(+) T cells. The results suggest that alloimmunization offers an alternative or complementary strategy in inducing an innate anti-viral factor that inhibits HIV-1 infection.
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PMID:The effect of allogeneic in vitro stimulation and in vivo immunization on memory CD4(+) T-cell APOBEC3G expression and HIV-1 infectivity. 1958 16

Mammary epithelial cells (MECs) are characterized by specific spatial architecture with several distinguishing features such as: polarized morphology, specialized cell-cell contacts, and attachment to an underlying basement membrane. Three dimensional (3D) basement membrane cultures provide a unique opportunity to model the architecture of epithelium in vitro. The aim of this study was to characterize the growth of bovine mammary epithelial cell line BME-UV1 in 3D culture on Matrigel and identification of differently expressed genes in bovine MECs forming polarized structures in comparison to conventional monolayer (2D) cell culture. We demonstrate that BME-UV1 cells grown on Matrigel form polarized acinar structures during 16 days of culture. A microarray study has proven that the difference in spatial architecture between MECs cultured in monolayer and 3D system is reflected by differences in transcriptomic profile. Microarray data analysis showed 40 differentially expressed genes with statistical significance (p<0.05) and characterized biological functions. Identified genes comprised of cytoskeletal proteins, extracellular matrix components, kinases such as: Rac serine/threonine kinase, SRPK, protooncogene tyrosine-protein kinase ABL1, uridine cytidine kinase and proteins with nucleic acid binding / transcription factor activity. Products of those genes are involved in processes which are known to participate in regulating mammary gland polarization and function.
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PMID:Differences in growth and transcriptomic profile of bovine mammary epithelial monolayer and three-dimensional cell cultures. 1960 9

Chronic lymphocytic leukaemia (CLL) is a heterogeneous disease exhibiting variable clinical course and survival rates. Mutational status of the immunoglobulin heavy chain variable regions (IGHVs) of CLL cells offers useful prognostic information for high-risk patients, but time and economical costs originally prevented it from being routinely used in a clinical setting. Instead, alternative markers of IGHV status, such as zeta-associated protein (ZAP70) or messenger RNA levels are often used. We report a (1)H-NMR-based metabolomics approach to examine serum metabolic profiles of early stage, untreated CLL patients (Binet stage A) classified on the basis of IGHV mutational status or ZAP70. Metabolic profiles of CLL patients (n=29) exhibited higher concentrations of pyruvate and glutamate and decreased concentrations of isoleucine compared with controls (n=9). Differences in metabolic profiles between unmutated (UM-IGHV; n=10) and mutated IGHV (M-IGHV; n=19) patients were determined using partial least square discriminatory analysis (PLS-DA; R(2)=0.74, Q(2)=0.36). The UM-IGHV patients had elevated levels of cholesterol, lactate, uridine and fumarate, and decreased levels of pyridoxine, glycerol, 3-hydroxybutyrate and methionine concentrations. The PLS-DA models derived from ZAP70 classifications showed comparatively poor goodness-of-fit values, suggesting that IGHV mutational status correlates better with disease-related metabolic profiles. Our results highlight the usefulness of (1)H-NMR-based metabolomics as a potential non-invasive prognostic tool for identifying CLL disease-state biomarkers.
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PMID:Serum metabolome analysis by 1H-NMR reveals differences between chronic lymphocytic leukaemia molecular subgroups. 2009 Jul 81


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