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:2.7.7.6 (RNA polymerase)
34,946 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Interphase chromatin is arranged into topologically separated domains comprising gene expression and replication units through genomic sequence elements, so-called MAR or SAR regions (for matrix- or scaffold-associating regions). S/MAR regions are located near the boundaries of actively transcribed genes and were shown to influence their activity. We show that scaffold attachment factor B (SAF-B), which specifically binds to S/MAR regions, interacts with RNA polymerase II (RNA pol II) and a subset of serine-/arginine-rich RNA processing factors (SR proteins). SAF-B localized to the nucleus in a speckled pattern that coincided with the distribution of the SR protein SC35. Furthermore, we show that overexpressed SAF-B induced an increase of the 10S splice product using an E1A reporter gene and repressed the activity of an S/MAR flanked CAT reporter gene construct in vivo . This indicates an association of SAF-B with SR proteins and components of the transcription machinery. Our results describe the coupling of a chromatin organizing S/MAR element with transcription and pre-mRNA processing components and we propose that SAF-B serves as a molecular base to assemble a 'transcriptosome complex' in the vicinity of actively transcribed genes.
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PMID:SAF-B protein couples transcription and pre-mRNA splicing to SAR/MAR elements. 967 16

The M1 protein of influenza virus inhibits the in vitro transcriptase activity of ribonucleoprotein cores from virions. This inhibitory activity is thought to be relevant in vivo because accumulation of M1 at the late stages of viral replication may be the cue to halt viral mRNA production. A model influenza reporter genome was used to explore the effect of M1 on the activity of the influenza virus transcriptase complex within cultured cells. Expression of M1 in cells bearing the model influenza virus reporter genome was accompanied by a reduction of CAT gene expression to 12% of control levels. Quantification of RNA by ribonuclease protection assay revealed that the influenza reporter genome mRNA levels in M1-expressing cells were reduced by approximately 74% compared with those of cells expressing a control protein. These findings are consistent with the proposed model in which M1 is responsible for limiting viral transcription during late stages of infection. By expressing truncated forms of M1, the inhibitory activity was found to reside within the amino-terminal half of the M1 protein. Two independent inhibitory domains were identified in this region: one between amino acid residues 1-90 and the other spanning residues 91-127.
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PMID:The matrix 1 protein of influenza A virus inhibits the transcriptase activity of a model influenza reporter genome in vivo. 974 Jul 76

We describe here a novel approach for detecting and quantitating human respiratory syncytial virus (RSV) based on expression of a reporter gene from an RSV minigenome. BHK cells were cytoplasmically transformed with a noncytopathic Sindbis virus replicon expressing T7 RNA polymerase. These cells were then cotransfected with T7 expression plasmids that contain the cDNA of an RSV minigenome and the genes for RSV nucleocapsid proteins N, P, and L. The minigenome contains a reporter gene such as lacZ or CAT flanked by cis-acting RSV transcription signals. Subsequent infection of these cells with RSV resulted in a high level of reporter gene expression which could be inhibited by ribavirin. Mock-infected cells exhibited background levels of expression. This assay can be used to quantitate RSV and titer neutralizing antibody and may be a valuable tool for screening compounds for anti-RSV activity. It serves as a prototype for other negative-strand RNA viruses.
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PMID:Detection and quantitation of human respiratory syncytial virus (RSV) using minigenome cDNA and a Sindbis virus replicon: a prototype assay for negative-strand RNA viruses. 981 15

Multidrug-resistant Mycobacterium tuberculosis infection is now world wide health problem. However, according to the recent advances of molecular biological technics, some of the genetic mechanisms of drug-resistance of M. tuberculosis has been uncovered. Generally, drug-resistance of M. tuberculosis was caused by point mutations in chromosomal gene. In isoniazid (INH) resistant M. tuberculosis, mutations and genetic deletions in catalase-peroxidase gene (katG), inhA gene, or alkyl hydroperoxide reductase gene were reported. We also found that about 15% of INH-resistant M. tuberculosis isolates lacked katG gene, and these isolates showed highly resistance to INH with MIC > or = 64 micrograms/ml. On the other hand, mutations and other genetic alterations in RNA polymerase beta subunit gene (rpoB) were the major mechanisms of resistance to rifampicin (RFP) with high frequencies of 90% or more. Our evaluation of the relationship between RFP susceptibility and genetic alteration in rpoB gene also showed that 95% of RFP-resistant M. tuberculosis isolates involved genetic alterations in 69 bp core region of rpoB gene. Moreover, these genetic alterations in rpoB gene were suspected as the resistant mechanism to other rifamycin antituberculosis drugs, such as rifabutin and KRM-1648. In addition, it was reported that point mutations in 16S rRNA gene (rrs) and ribosomal protein S12 gene (rpsL) induced M. tuberculosis as streptomycin (SM) resistant phenotype. We analyzed genetic alternations in rpsL gene of clinically isolates of M. tuberculosis, about 60% of SM resistant isolates were shown point mutation in this gene ant they were all high SM-resistant with MIC > or = 256 micrograms/ml. Furthermore, nicotinamidase (pncA) gene, DNA gyrase A subunit (gyrA) gene, and embB gene were reported as the responsible gene to pyrazinamide-, quinolone- and ethambutol-resistance, respectively. Although all mechanisms of drug-resistance were still unclear, these informations are very useful and helpful for development of rapid diagnosis system of drug-resistant M. tuberculosis.
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PMID:[Multidrug-resistant tuberculosis. 2. Mechanisms of drug-resistance in Mycobacterium tuberculosis--genetic mechanisms of drug-resistance]. 986 28

We report here the in vitro characterization of PrpoB-345, the tobacco rpoB promoter recognized by NEP, the phage-type plastid RNA polymerase. Transcription extracts were prepared from mutant tobacco plants lacking PEP, the Escherichia coli-like plastid-encoded RNA polymerase. Systematic dissection of a approximately 1 kb fragment determined that the rpoB promoter is contained in a 15-nucleotide segment (-14 to +1) upstream of the transcription initiation site (+1). Point mutations at every nucleotide reduced transcription, except at the -5 position which was neutral. Critical for rpoB promoter function was a CRT-motif (CAT or CGT) at -8 to -6 (transcription <30%), defining it as the promoter core. The core CAT sequence is also present in the maize rpoB promoter, which is faithfully recognized by tobacco extracts. Alignment of NEP promoters identified a CATA or TATA (=YATA) sequence at the rpoB core position, also present in plant mitochondrial promoters. Furthermore, NEP and the phage T7 RNA polymerase exhibit similar sensitivity to inhibitors of transcription. These data indicate that the nuclear RpoZ gene, identified by sequence conservation with mitochondrial RNA polymerases, encodes the NEP catalytic subunit.
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PMID:In vitro characterization of the tobacco rpoB promoter reveals a core sequence motif conserved between phage-type plastid and plant mitochondrial promoters. 987 67

According to the recent advances of molecular biological technics, some of the genetic mechanisms of drug-resistance of Mycobacteria has been uncovered. Generally, drug-resistance of Mycobacterium tuberculosis was caused by point mutations in chromosomal gene. In isoniazid (INH) resistant M. tuberculosis, mutations and genetic deletions in catalase-peroxidase gene (katG), inhA gene, or alkyl hydroperoxide reductase gene were reported. On the other hand, mutations and other genetic alterations in RNA polymerase beta subunit gene (rpoB) were the major mechanisms of resistance to rifampicin (RFP) with high frequencies of 90% or more. Moreover, these genetic alterations in rpoB gene were suspected as the resistant mechanism to other rifamycin antituberculosis drugs, such as rifabutin. In addition, it was reported that point mutations in 16S rRNA gene (rrs) and ribosomal protein S12 gene (rpsL) induced M. tuberculosis as streptomycin (SM) resistant phenotype. Furthermore, nicotinamidase (pncA) gene, DNA gyrase A subunit (gyrA) gene, and embB gene were reported as the responsible gene to pyrazinamide-, quinolone- and ethambutol-resistance, respectively. Although all mechanisms of drug-resistance were still unclear, these information are very useful and helpful for development of rapid diagnosis system of drug-resistant M. tuberculosis.
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PMID:[Mechanisms of drug-resistance in mycobacteria]. 988 3

The prenatal diagnosis of peroxisomal D-3-hydroxyacyl-coenzyme A (CoA) dehydratase/D-3-hydroxyacyl-CoA dehydrogenase bifunctional protein (D-BP) deficiency was performed by peroxisomal beta-oxidation assay, indirect immunofluorescence staining, immunoblot analysis, and gene analysis of cultured amniocytes obtained from a fetus at 16 weeks' gestational age. beta-Oxidation activity, measured by [1-14C] lignoceric acid oxidation, was markedly decreased compared with the controls. Large peroxisomes were readily identified by immunofluorescence staining with anti-human catalase, as was found in the reported patients. Immunoreactive D-BP material was absent on immunoblot analysis and immunofluorescence staining with anti-human D-BP. Reverse transcriptase polymerase chain reaction (RT-PCR) analysis revealed the presence of the same 237-bp deletion in the cDNA as that detected in a sibling (the proband). The autopsied fetus showed the characteristic facial appearance and D-BP was deficient on immunoblot and immunohistopathological studies of the fetal tissues. No neuronal migration disorder was identified. This seems to be the first prenatal diagnosis of D-BP deficiency.
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PMID:Prenatal diagnosis of peroxisomal D-3-hydroxyacyl-CoA dehydratase/D-3-hydroxyacyl-CoA dehydrogenase bifunctional protein deficiency. 1031 76

Measles virus encodes three proteins required for the encapsidation, transcription and replication of viral genomes. The genes for these proteins have been inserted into the vaccinia virus genome together with the gene for the bacteriophage T7 RNA polymerase. Cells infected with this recombinant virus were able to encapsidate, transcribe and replicate a CAT gene positioned in the negative polarity behind a T7 promoter and flanked by measles virus genomic termini. Inhibition of the accumulation of the nucleocapsid proteins by actinomycin D led to an increase in CAT expression. Thus the measles virus polymerase activity, encoded by the vaccinia genome, was regulated by the level of measles proteins just as the authentic polymerase. The recombinant vaccinia described in this study could be useful for the production of measles virus-like particles encoding foreign genes and employed in vaccination or gene therapy strategies.
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PMID:A functional measles virus replication and transcription machinery encoded by the vaccinia virus genome. 1032 36

We have cloned a 3.6-kb genomic DNA fragment from Pseudomonas aeruginosa harboring the rpoA, rplQ, katA, and bfrA genes. These loci are predicted to encode, respectively, (i) the alpha subunit of RNA polymerase; (ii) the L17 ribosomal protein; (iii) the major catalase, KatA; and (iv) one of two iron storage proteins called bacterioferritin A (BfrA; cytochrome b1 or b557). Our goal was to determine the contributions of KatA and BfrA to the resistance of P. aeruginosa to hydrogen peroxide (H2O2). When provided on a multicopy plasmid, the P. aeruginosa katA gene complemented a catalase-deficient strain of Escherichia coli. The katA gene was found to contain two translational start codons encoding a heteromultimer of approximately 160 to 170 kDa and having an apparent Km for H2O2 of 44.7 mM. Isogenic katA and bfrA mutants were hypersusceptible to H2O2, while a katA bfrA double mutant demonstrated the greatest sensitivity. The katA and katA bfrA mutants possessed no detectable catalase activity. Interestingly, a bfrA mutant expressed only approximately 47% the KatA activity of wild-type organisms, despite possessing wild-type katA transcription and translation. Plasmids harboring bfrA genes encoding BfrA altered at critical amino acids essential for ferroxidase activity could not restore wild-type catalase activity in the bfrA mutant. RNase protection assays revealed that katA and bfrA are on different transcripts, the levels of which are increased by both iron and H2O2. Mass spectrometry analysis of whole cells revealed no significant difference in total cellular iron levels in the bfrA, katA, and katA bfrA mutants relative to wild-type bacteria. Our results suggest that P. aeruginosa BfrA may be required as one source of iron for the heme prosthetic group of KatA and thus for protection against H2O2.
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PMID:Bacterioferritin A modulates catalase A (KatA) activity and resistance to hydrogen peroxide in Pseudomonas aeruginosa. 1036 48

Transforming growth factor (TGF)-beta1 is a growth factor involved in the mechanisms of lung repair and fibrosis that follow inflammatory processes. We sought to examine the link between the generation of reactive oxygen intermediates (ROI) or reactive nitrogen intermediates (RNI) by inflammatory cells and the expression of TGF-beta1 by alveolar epithelial cells. Exposure of the A549 lung epithelial cell line to either an ROI generating system (xanthine and xanthine oxidase) or an RNI donor (S-nitroso-N-acetyl-penicillamine [SNAP]) promoted a time- and dose-dependent increase in TGF-beta1 release, as measured by a specific enzyme-linked immunosorbent assay. At the peak, the levels of TGF-beta1 were twice the control values. The induction of TGF-beta1 release by ROI was blunted by catalase and unaffected by superoxide dismutase, indicating the involvement of hydrogen peroxide. The response was also blunted by 5, 6-dichloro-1-beta-D-ribofuranosyl benzimidazole (DRB), a specific RNA polymerase II inhibitor, and accompanied by a corresponding increase in TGF-beta1 messenger RNA, as measured by quantitative/competitive reverse transcription polymerase chain reaction, suggesting the involvement of transcriptional mechanisms and possibly other downstream mechanisms. In contrast, RNI-induced TGF-beta1 release was unaffected by DRB and blunted by the protein synthesis inhibitor cycloheximide, suggesting the involvement of translational and post-translational mechanisms. This response required cyclic guanosine monophosphate (cGMP)- mediated processes because (1) immunoreactive cGMP accumulated in the culture medium of SNAP-treated cells; (2) SNAP-induced TGF-beta1 release was blunted by KT 5823, an inhibitor of cGMP-dependent protein kinase; and (3) similar increase in TGF-beta1 release was obtained by cell exposure to membrane-permeable dibutyryl-cGMP or to atrial natriuretic factor, a known agonist of particulate guanylate cyclase. These data suggest that in vitro exposure of human alveolar epithelial cells to ROI and RNI enhances TGF-beta1 release through different mechanisms. In vivo, this control may constitute a molecular link between inflammatory and fibrotic processes.
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PMID:Reactive oxygen and nitrogen intermediates increase transforming growth factor-beta1 release from human epithelial alveolar cells through two different mechanisms. 1038 1


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