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Site Upstream (site + upstream)

Distribution by Scientific Domains
Life Sciences66%
Medical Sciences22%

Kinds of Site Upstream

  • binding site upstream
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    Selected Abstracts

    Regulation of anaerobic arginine catabolism in Bacillus licheniformis by a protein of the Crp/Fnr family

    FEMS MICROBIOLOGY LETTERS, Issue 2 2000
    Abdelouahid Maghnouj
    Abstract Arginine anaerobic catabolism occurs in Bacillus licheniformis through the arginine deiminase pathway, encoded by the gene cluster arcABDC. We report here the involvement of a new protein, ArcR, in the regulation of the pathway. ArcR is a protein of the Crp/Fnr family encoded by a gene located 109 bp downstream from arcC. It binds to a palindromic sequence, very similar to an Escherichia coli Crp binding site, located upstream from arcA. Residues in the C-terminal domain of Crp that form the DNA binding motif, in particular residues Arg-180 and Glu-181 that make specific bonds with DNA, are conserved in ArcR, suggesting that the complexes formed with DNA by Crp and ArcR are similar. Moreover, the pattern of DNase I hypersensitivity sites induced by the binding of ArcR suggests that ArcR bends the DNA in the same way as Crp. From the absence of anaerobic induction following inactivation of arcR and from the existence of a binding site upstream of the arcA transcription start point, it can be inferred that ArcR is an activator of the arginine deiminase pathway. [source]

    Reduced levels of miR-34a in neuroblastoma are not caused by mutations in the TP53 binding site,

    GENES, CHROMOSOMES AND CANCER, Issue 7 2009
    Galina Feinberg-Gorenshtein
    Neuroblastoma (NB) is the most common extracranial solid tumor in children below the age of 5 years. miR-34a, located in chromosome band 1p36, has been recently implicated as a tumor suppressor gene in NB. In addition, it has been shown that miR-34a is activated by TP53 by binding to a TP53 binding site upstream to the mature miR-34a. We studied NB tumors from 57 patients for miR-34a expression levels, 1p status, mutations in the TP53 coding region and mutations of the TP53 binding site. Reduced expression levels of miR-34a were identified in tumors harboring 1p36.3 Loss (P = 0.028). No mutations were identified in the coding region of TP53, or in the TP53 binding site. Thus, mutations in the binding site are not an additional mechanism for the inactivation of miR-34a in NB. Other regulatory mechanisms controlling miR-34a expression and its relationship to TP53 should be further explored. © 2009 Wiley-Liss, Inc. [source]

    The cia operon of Streptococcus mutans encodes a unique component required for calcium-mediated autoregulation

    MOLECULAR MICROBIOLOGY, Issue 1 2008
    Xuesong He
    Summary Streptococcus mutans is a primary pathogen for dental caries in humans. CiaR and CiaH of S. mutans comprise a two-component signal transduction system (TCS) involved in regulating various virulent factors. However, the signal that triggers the CiaRH response remains unknown. In this study, we show that calcium is a signal for regulation of the ciaRH operon, and that a double-glycine-containing small peptide encoded within the ciaRH operon (renamed ciaX) mediates this regulation. CiaX contains a serine + aspartate (SD) domain that is shared by calcium-binding proteins. A markerless in-frame deletion of ciaX reduced ciaRH operon expression and diminished the calcium repression of operon transcription. Point mutations of the SD domain resulted in the same phenotype as the in-frame deletion, indicating that the SD domain is required for CiaX function. Further characterization of ciaX demonstrated that it is involved in calcium-mediated biofilm formation. Furthermore, inactivation of ciaR or ciaH led to the same phenotype as the in-frame deletion of ciaX, suggesting that all three genes are involved in the same regulatory pathway. Sequence analysis and real-time RT-PCR identified a putative CiaR binding site upstream of ciaX. We conclude that the ciaXRH operon is a three-component, self-regulatory system modulating cellular functions in response to calcium. [source]

    Submicromolar hydrogen peroxide disrupts the ability of Fur protein to control free-iron levels in Escherichia coli

    MOLECULAR MICROBIOLOGY, Issue 3 2007
    Shery Varghese
    Summary In aerobic environments, mutants of Escherichia coli that lack peroxidase and catalase activities (Hpx,) accumulate submicromolar concentrations of intracellular H2O2. We observed that in defined medium these strains constitutively expressed members of the Fur regulon. Iron-import proteins, which Fur normally represses, were fully induced. H2O2 may antagonize Fur function by oxidizing the Fur:Fe2+ complex and inactivating its repressor function. This is a potential problem, as in iron-rich environments excessive iron uptake would endanger H2O2 -stressed cells by accelerating hydroxyl-radical production through the Fenton reaction. However, the OxyR H2O2 -response system restored Fur repression in iron-replete Luria,Bertani medium by upregulating the synthesis of Fur protein. Indeed, when the OxyR binding site upstream of fur was disrupted, Hpx, mutants failed to repress transporter synthesis, and they exhibited high levels of intracellular free iron. Mutagenesis and bacteriostasis resulted. These defects were eliminated by mutations or chelators that slowed iron import, confirming that dysregulation of iron uptake was the root problem. Thus, aerobic organisms must grapple with a conundrum: how to monitor iron levels in oxidizing environments that might perturb the valence of the analyte. The induction of Fur synthesis by the OxyR response comprises one evolutionary solution to that problem. [source]

    Reconstituting retroviral (ReCon) vectors facilitating delivery of cytotoxic genes in cancer gene therapy approaches

    THE JOURNAL OF GENE MEDICINE, Issue 2 2008
    Eva Maria Brandtner
    Abstract Background We have previously described the generation of reconstituting retroviral (ReCon) vectors designed for cancer gene therapy using cytotoxic gene products. The unique vector structure with a promoter physically separated from the transgene allows generation of stable virus producer cells irrespective of the toxic gene. The mechanism of synthesis of DNA from retroviral RNA dictates that infection leads to the reconstitution of functional expression cassettes in the target cell. Methods To improve vector titres, a cytomegalovirus enhancer was inserted upstream of the 5,-long-terminal repeat (LTR); the Woodchuck hepatitis virus post-transcriptional regulatory element and an elongated attachment site upstream of the 3,-LTR were included. In addition, a bacterial origin of replication was deleted and a functional internal polyadenylation signal mutated. Transcriptional targeting was attempted by introducing mammary tissue-specific promoters such as the U3 region of mouse mammary tumour virus or the promoter of the whey acidic protein encoding gene. All modifications were analysed in detail with respect to virus production and infectivity. Finally, the vector was armed with the ,-holin encoding gene and transduced cells were analysed for cytotoxic effects. Results Distinct modifications of the vector resulted in a titre improvement of more than 560-fold. Compatibility of the optimized vector with targeted cellular promoters was demonstrated. When equipped with the cytotoxic gene, stable producer cells could be successfully established and high titre virus infection resulted in rigorous target cell killing. Conclusions The ReCon vector in its optimized form is an attractive tool for cancer gene therapy approaches. Copyright © 2007 John Wiley & Sons, Ltd. [source]

    Real-time observation of Wnt ,-catenin signaling in the chick embryo

    DEVELOPMENTAL DYNAMICS, Issue 1 2010
    Anne C. Rios
    Abstract A critical mediator of cell,cell signaling events during embryogenesis is the highly conserved Wnt family of secreted proteins. Reporter constructs containing multimerized TCF DNA binding sites have been used to detect Wnt ,-catenin dependent activity during animal development. In this report, we have constructed and compared several TCF green fluorescent protein (GFP) reporter constructs. They contained 3, 8, or 12 TCF binding sites upstream of a minimal promoter driving native or destabilized enhanced GFP (EGFP). We have used the electroporation of somites in the chick embryo as a paradigm to test them in vivo. We have verified that they all respond to Wnt signaling in vivo. We have then assessed their efficiency at reflecting the activity of the Wnt pathway. Using destabilized EGFP reporter constructs, we show that somite cells dynamically regulate Wnt/,-catenin,dependent signaling, a finding that was confirmed by performing time-lapse video confocal observation of electroporated embryos. Developmental Dynamics 239:346,353, 2010. © 2009 Wiley-Liss, Inc. [source]

    Biomarker study of a municipal effluent dispersion plume in two species of freshwater mussels

    ENVIRONMENTAL TOXICOLOGY, Issue 3 2002
    F. Gagné
    Abstract The toxicological effects of a primary-treated municipal effluent plume were investigated in two species of freshwater mussels, Elliptio complanata and Dreissena polymorpha, exposed for 62 days at sites upstream and downstream of an effluent outfall in the St. Lawrence River (Quebec, Canada). Levels of metallothioneins (MT), cytochrome P4501A1 activity, DNA damage, total lipids, relative levels of vitellins, and phagocytic activity (in E. complanata hemocytes) were determined after the exposure period. A parallel analysis measured heavy metals and coprostanol in mussel tissues. The results show that significant levels of coprostanol and some metals (specifically, Cu, Hg, Sb, Se, and Zn) had accumulated in mussels caged 5 km downstream of the effluent plume. Mixed-function oxidase activity, MT in gills, total lipids, DNA damage (in D. polymorpha only), and total hemolymph bacteria (in E. complanata only) had increased in these mussels, while levels of total cadmium (Cd), MT in digestive glands or whole soft tissues, phagocytic activity, and DNA damage in the digestive gland (in E. complanata only) were diminished. The exposure of mussels to surface waters contaminated by a municipal effluent led to many stress responses, depending on both the tissues and the species being examined. © 2002 Wiley Periodicals, Inc. Environ Toxicol 17: 149,159, 2002; Published online in Wiley InterScience (www.interscience.wiley.com). DOI 10.1002/tox.10046 [source]

    Impacts of aircraft deicer and anti-icer runoff on receiving waters from Dallas/Fort worth International Airport, Texas, USA

    ENVIRONMENTAL TOXICOLOGY & CHEMISTRY, Issue 11 2006
    Steven R. Corsi
    Abstract From October 2002 to April 2004, data were collected from Dallas/Fort Worth (DFW) International Airport (TX, USA) outfalls and receiving waters (Trigg Lake and Big Bear Creek) to document the magnitude and potential effects of aircraft deicer and anti-icer fluid (ADAF) runoff on water quality. Glycol concentrations at outfalls ranged from less than 18 to 23,800 mg/L, whereas concentrations in Big Bear Creek were less because of dilution, dispersion, and degradation, ranging from less than 18 to 230 mg/L. Annual loading results indicate that 10 and 35% of what was applied to aircraft was discharged to Big Bear Creek in 2003 and 2004, respectively. Glycol that entered Trigg Lake was diluted and degraded before reaching the lake outlet. Dissolved oxygen (DO) concentrations at airport outfalls sometimes were low (<2.0 mg/L) but typical of what was measured in an urban reference stream. In comparison, the DO concentration at Trigg Lake monitoring sites was consistently greater than 5.5 mg/L during the monitoring period, probably because of the installation of aerators in the lake by DFW personnel. The DO concentration in Big Bear Creek was very similar at sites upstream and downstream of airport influence (>5.0 mg/L). Results of toxicity tests indicate that effects on Ceriodaphnia dubia, Pimephales promelas, and Selanastrum capricornutum are influenced by type IV ADAF (anti-icer), not just type I ADAF (deicer) as is more commonly assumed. [source]

    DNA hypomethylation in rheumatoid arthritis synovial fibroblasts

    ARTHRITIS & RHEUMATISM, Issue 12 2009
    Emmanuel Karouzakis
    Objective Rheumatoid arthritis synovial fibroblasts (RASFs) are phenotypically activated and aggressive. We undertook this study to investigate whether the intrinsic activation of RASFs is due to global genomic hypomethylation, an epigenetic modification. Methods Global genomic hypomethylation was assessed by immunohistochemistry, flow cytometry, and L1 promoter bisulfite sequencing. The levels of Dnmt1 were determined in synovial tissue and cultured SFs by Western blotting before and after treatment with cytokines and growth factors. Normal SFs were treated for 3 months with a nontoxic dose of the DNA hypomethylation drug 5-azacytidine (5-azaC), and changes in gene expression were revealed using complementary DNA arrays. The phenotypic changes were confirmed by flow cytometry. Results In situ and in vitro, RASF DNA had fewer 5-methylcytosine and methylated CG sites upstream of an L1 open-reading frame than did DNA of osteoarthritis SFs, and proliferating RASFs were deficient in Dnmt1. Using 5-azaC, we reproduced the activated phenotype of RASFs in normal SFs. One hundred eighty-six genes were up-regulated >2-fold by hypomethylation, with enhanced protein expression. These included growth factors and receptors, extracellular matrix proteins, adhesion molecules, and matrix-degrading enzymes. The hypomethylating milieu induced irreversible phenotypic changes in normal SFs, which resembled those of the activated phenotype of RASFs. Conclusion DNA hypomethylation contributes to the chronicity of RA and could be responsible for the limitation of current therapies. [source]