Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.14.14.3 (luciferase)
 38,195 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The newly described iron transporter, ferroportin (MTP1, IREG1), is expressed in a variety of tissues including the duodenum and cells of the mononuclear phagocyte system (MPS). In the MPS, ferroportin is hypothesized to be a major exporter of iron scavenged from senescent erythrocytes. Changes in iron metabolism, including the sequestration of iron in the MPS, are characteristic of both acute and chronic inflammation and these conditions induce changes in ferroportin expression. In a mouse model of acute inflammation, LPS administration is associated with reduced MPS ferroportin protein and mRNA expression. In addition, the ferroportin 5' UTR also has an iron-responsive element that binds to the iron-response proteins, but whether there is a role for this IRE in inflammation induced regulation of ferroportin has been unclear. A luciferase reporter gene under the control of the mouse ferroportin promoter and 5' UTR was used to determine if this 5' UTR conferred IRE-dependent regulation on this reporter gene. Stimulation of reporter gene transfected RAW 264.7 cells (a mouse macrophage cell line) with LPS resulted in IRE-dependent inhibition of luciferase production. Inhibitors of nitric oxide synthase abrogated the IRE-dependent effect of LPS. In addition, direct treatment of RAW 264.7 and with NO donor S-nitroso-N-acetylpenicillamine resulted in IRE-dependent down-regulation of luciferase expression. The effect of NO was consistent with IRP1/IRE mediated translation block. There are most likely both inflammation-mediated transcriptional and post-transcriptional (IRE-dependent) mechanisms for inhibiting ferroportin expression in MPS cells.
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PMID:Role of the ferroportin iron-responsive element in iron and nitric oxide dependent gene regulation. 1254 22

X-linked sideroblastic anemia (XLSA) is caused by mutations in the erythroid-specific 5-aminolevulinate synthase gene (ALAS2). XLSA was diagnosed in a 32-year-old woman with a mild phenotype and moderately late onset. Pyridoxine therapy had no effect in the proband, but in her affected son engendered a modest increase in hemoglobin concentration and a 4-fold reduction in ferritin iron. Molecular analysis identified a C to G transversion at nucleotide -206 from the transcription start site, as defined by primer extension, in the proximal promoter region of ALAS2. No other mutations were found in the promoter region, the flanking intronic sequences, the exons, or the 3' genomic region. The same mutation was found in her affected son but not in any other of her unaffected relatives. The mutation resulted in a 94% loss of activity relative to the wild-type sequence for a luciferase reporter construct containing the proximal 293 nucleotides (nt's) of the ALAS2 promoter when transfected into human erythroid K562 cells. Confirming the mutation's deleterious effect, the ALAS2 mRNA level in the proband's erythroid precursors was reduced 87%. The mutation occurred in or near 3 different putative transcription factor binding sites of unknown erythroid importance. The dramatic decreases in reporter activity and mRNA level suggest that the region of the mutation may bind a novel and important erythroid regulatory element.
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PMID:A promoter mutation in the erythroid-specific 5-aminolevulinate synthase (ALAS2) gene causes X-linked sideroblastic anemia. 1266 58

Gene delivery using cationic liposomes results in relatively low transfection, especially under in vivo conditions. This system, however, can overcome some of the problems associated with viral delivery systems. The present study was carried out in order to improve the transfection efficiency of cationic liposomes by preparing magnetic cationic liposomes (MCL). Small MCL approximately 40 nm in diameter and incorporating one or two magnetite particles were prepared with phosphatidylethanolamine and 3beta-[N-(N', N'-dimethylaminoethane)-carbamoyl] cholesterol. The efficiency of MCL in gene delivery was evaluated by using plasmid DNA containing a luciferase reporter gene and human osteosarcoma Saos-2 cells. Without a magnetic field, maximum luciferase activity was observed when DNA was mixed with MCL at a 1:5 ratio and incubated with cells for 6 h. Under a magnetic field, maximum luciferase activity was achieved by 30-min magnetic induction. This improvement in transfection efficiency by magnetic induction was approximately 3.5-fold. The feasibility of this active transgenic system was further shown by measuring apoptosis rates after transfection of the p53 gene to Saos-2 cells. Apoptosis rates increased to 18.9% from 2.4% by magnetic induction. In conclusion, a gene delivery system including MCL and magnetic induction was found to achieve rapid and enhanced gene delivery in vitro. Such a gene delivery system may be applicable under in vivo conditions, and is expected to offer numerous clinical advantages.
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PMID:Targeted gene delivery to human osteosarcoma cells with magnetic cationic liposomes under a magnetic field. 1268 73

Iron (Fe) chelators induce a G1/S arrest and several of these are undergoing clinical trials as anticancer agents. Despite this, little is known concerning the precise function of Fe in cell cycle progression and the role of p53 in this process. The aim of this study was to assess the effect of Fe chelators on p53 and the mechanism involved in the chelator-mediated increase in mRNA levels of the universal cyclin-dependent kinase inhibitor p21CIP1/WAF1. Cells were incubated with the potent Fe chelator 2-hydroxy-1-naphthylaldehyde isonicotinoyl hydrazone (311) and the results compared with those from cells treated with actinomycin D (Act D), which induces p53. Following incubation with 311, a 3- to 5-fold increase in nuclear p53 protein was observed in cells with wild-type p53. In addition, 311 increased p53 DNA-binding activity 2-fold, while Act D increased it 3- to 5-fold in cells with native p53. To determine the role of p53 in WAF1 transcription, a reporter construct was used consisting of a WAF1 promoter containing the p53-binding site. In cells with wild-type p53, chelators had no effect on luciferase activity, while the positive control, Act D, caused a significant increase. Hence, despite increased p53 protein expression and p53 DNA-binding activity following chelation, these latter results suggested it had no role in up-regulating WAF1 mRNA. Our experiments demonstrated: (i) that the elevated WAF1 mRNA expression after Fe chelation was due to increased transcription and also to a post-transcriptional mechanism that was sensitive to cycloheximide; and (ii) that Fe-chelation increased WAF1 expression through a p53-independent pathway.
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PMID:The effect of potent iron chelators on the regulation of p53: examination of the expression, localization and DNA-binding activity of p53 and the transactivation of WAF1. 1286 19

We screened for drugs that specifically interact with the 5'-untranslated region of the mRNA encoding the Alzheimer's amyloid precursor protein (APP). Our goal was to use newly discovered APP 5' UTR directed compounds to limit amyloid-beta (Abeta)-peptide output in cell culture systems. The APP 5' UTR folds into a stable RNA secondary structure (Gibbs free energy: DeltaG = -54.9 kcal/mol) and is an important regulator of the amount of APP translated in response to IL-1 (Nilsson et al., 1998; Rogers et al., 1999) and iron (Rogers et al., 2002). Seventeen drug "hits" were identified from a library of 1,200 FDA preapproved drugs (Rogers et al., 2002). Six of the original 17 compounds were validated for their capacity to suppress reporter gene expression in stable neuroblastoma transfectants expressing the dicistronic reporter construct shown in Fig. 2. These six leads suppressed APP 5' UTR driven luciferase translation while causing no effect on the translation of dicistronic GFP gene translated from a viral IRES (negative control to ensure specificity during drug screens). In this report, we show that paroxetine (serotonin reuptake blocker) and dimercaptopropanol (Hg chelator) exerted significant effects on APP expression (steady-state levels of APP), whereas Azithromycin altered APP processing. None of these three compounds altered APLP-1 expression. In the future, we will identify further novel compounds that influence Abeta levels, either via translation inhibition or by changing the activity of proteins coupled between APP translation and APP processing.
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PMID:Drug discovery targeted to the Alzheimer's APP mRNA 5'-untranslated region: the action of paroxetine and dimercaptopropanol. 1450 Oct 7

Candida albicans is an opportunistic pathogen that has adapted uniquely to life in mammalian hosts. One of the host factors recognized by this yeast is hemoglobin, which binds to a specific cell surface receptor. In addition to its regulating the expression of adhesion receptors on the yeast, we have found that hemoglobin induces the expression of a C. albicans heme oxygenase (CaHmx1p). Hemoglobin transcriptionally induces the CaHMX1 gene independent of the presence of inorganic iron in the medium. A Renilla luciferase reporter driven by the CaHMX1 promoter demonstrated rapid activation of transcription by hemoglobin and (cobalt protoporphyrin IX) globin but not by apoglobin or other proteins. In contrast, iron deficiency or exogenous hemin did not activate the reporter until after 3 h, suggesting that induction of the promoter by hemoglobin is mediated by receptor signaling rather than heme or iron flux into the cell. As observed following disruption of the Saccharomyces cerevisiae ortholog, HMX1, a CaHMX1 null mutant was unable to grow under iron restriction. This suggests a role for CaHmx1p in inorganic iron acquisition. CaHMX1 encodes a functional heme oxygenase. Exogenous heme or hemoglobin is exclusively metabolized to alpha-biliverdin. CaHMX1 is required for utilization of these exogenous substrates, indicating that C. albicans heme oxygenase confers a nutritional advantage for growth in mammalian hosts.
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PMID:Heme oxygenase in Candida albicans is regulated by hemoglobin and is necessary for metabolism of exogenous heme and hemoglobin to alpha-biliverdin. 1461 78

Association of various autoimmune and infectious diseases with genetic variation in the solute carrier family 11 member 1 (SLC11A1) gene, formerly known as the natural resistance-associated macrophage protein 1 (NRAMP1) gene, is in accordance with its role in iron metabolism and immune function. In this investigation, in vitro studies were performed to determine whether allelic variants in the promoter region of the gene are affected by iron loading, thereby leading to differential expression of SLC11A1. Constructs containing five different SLC11A1 5'-(GT)n polymorphic alleles identified in the South African population (alleles 2, 3, 5, 8, and 9) and a C to T point mutation at nucleotide position -237, both in the absence and presence of allele 3, were cloned into the pGL2-Basic luciferase-reporter vector and transfected into U937 and THP-1 cells. Addition of exogenous stimuli, including interferon-gamma, bacterial lipopolysaccharide, and ferric ammonium citrate, demonstrated significant differences in the ability of these alleles to regulate gene expression. Striking differences were obtained upon iron loading, with allele 3 showing opposite effects in the presence or absence of promoter polymorphism -237C-->T. Our findings provide direct evidence that this promoter polymorphism is functional and support the hypothesis that iron dysregulation mediated by allelic effects of SLC11A1 underlies disease susceptibility linked to infectious and autoimmune conditions.
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PMID:Expression of the SLC11A1 (NRAMP1) 5'-(GT)n repeat: opposite effect in the presence of -237C-->T. 1522 10

Ferrochelatase (FECH), the last enzyme of the heme biosynthetic pathway, catalyzes the insertion of iron into protoporphyrin to form heme. This pathway provides heme for hemoglobin and other essential hemoproteins. The regulatory role of oxygen in the pathway has not been clearly established. In this study, we examined whether FECH gene expression is upregulated during hypoxia by a mechanism which involves the hypoxia-inducible factor 1 (HIF-1). Two HIF-1 binding motifs were identified within the -150 bp FECH minimal promoter sequence. Exposure of HEL, K562, and Hep-G2 cells to hypoxia for 18 hours resulted in a significant increase in FECH mRNA expression (p < 0.05). Hypoxia also transactivated the minimal promoter for the FECH gene in the cells. Transient co-expression of wild-type HIF-1alpha or a dominant negative HIF-1alpha with the FECH minimal promoter luciferase construct stimulated or blocked FECH promoter activity, respectively. Expression of the von Hippel-Lindau (VHL) tumor suppressor factor blocked the expression of both FECH mRNA and HIF-1alpha protein during normoxic culture of renal carcinoma cell line (RCC4). The results suggest that the FECH gene is a target for HIF-1 during hypoxia.
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PMID:Regulation of ferrochelatase gene expression by hypoxia. 1531 48

The 5' untranslated region (5'UTR) of the transcript encoding the Alzheimer's amyloid precursor protein (APP) is a key regulatory sequence that determines the amount of intracellular APP holoprotein present in brain derived cells. Using neuroblastoma cells (SY5Y) we developed a transfection based screen of a library of FDA drugs to identify compounds that limited APP luciferase reporter expression translated from the APP 5'UTR. Paroxetine (Paxil trade mark ), dimercaptopropanol, phenserine, desferrioxamine, tetrathiolmobdylate, and azithromycin were six leads that were subsequently found to also suppress APP holoprotein levels or to alter APP cleavage (azithromycin). Since APP holoprotein levels are proportionate to Abeta peptide output in many systems we tested the efficacy of paroxetine and dimercaptopropanol to limit Abeta secretion as measured by ELISA assays. Paroxetine and dimercaptopropanol limited Abeta peptide secretion from lens epithelial cells (B3 cells). Interestingly, paroxetine changed the steady-state levels of transferrin receptor mRNAs. These data suggested that this serotonin reuptake inhibitor (SSRI) provided extra pharmacological action to chelate interacellular iron or change the intracellular iron distribution. An altered iron distribution would be predicted to indirectly limit APP holoprotein expression and Abeta peptide secretion.
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PMID:FDA-preapproved drugs targeted to the translational regulation and processing of the amyloid precursor protein. 1531 61

Increased iron store in the body may increase the risk of many diseases such as cancer and inflammation. However, the precise pathogenic mechanism of iron has not yet been elucidated. In the present study, the early biological responses of cells to iron treatment were investigated in AP-1 luciferase reporter stably transfected mouse epidermal JB6 cells and primary rat hepatocytes. It was shown that water-soluble iron compounds, such as FeSO4 and Fe2(SO4)3, were more active in inducing AP-1 in JB6 cells than water-insoluble iron compounds, such as Fe2O3 and FeS. Iron stimulated mitogen-activated protein kinase (MAPK) family members of extracellular signal-regulated kinases (ERKs) and p38 MAPK but not c-jun NH2 terminal kinases (JNKs), both in JB6 cells and in primary rat hepatocytes, as determined by the phosphorylation assay. Interestingly, the increase in AP-1 luciferase activity by iron was inhibited by the pretreatment of the cells with PD98059, a specific MEK1 inhibitor, and SB202190, a p38 kinase inhibitor. Levels of interleukin-6 (IL-6), a pro-inflammatory cytokine, were increased in JB6 cells by iron in a dose-dependent manner. The increase in IL-6 and its mRNA by iron was also eliminated by the pretreatment of the cells with PD98059 and SB202190. Since the IL-6 promoter contains an AP-1 binding site, our studies indicate that the iron-induced IL-6 gene expression may be mediated through ERKs and p38 MAPK pathways, possibly one of the important mechanisms for the pathogenesis of iron overload.
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PMID:Iron-induced interleukin-6 gene expression: possible mediation through the extracellular signal-regulated kinase and p38 mitogen-activated protein kinase pathways. 1536 95


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