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)

RNA interference (RNAi) is implicated in maintaining tandem DNA arrays as constitutive heterochromatin. We used chromatin immunoprecipitation with antibodies to RNA polymerase II (RNAPol-ChIP) to test for transcription of the following repeat arrays in human cells: subtelomeric D4Z4, pericentromeric satellite 2, and centromeric satellite alpha. D4Z4 has a promoter-like sequence upstream of an ORF in its 3.3-kb repeat unit. A short D4Z4 array at 4q35 is linked to facioscapulohumeral muscular dystrophy (FSHD). By RNAPol-ChIP and RT-PCR, little or no transcription of D4Z4 was detected in FSHD and normal myoblasts; lymphoblasts from an FSHD patient, a control, and a patient with D4Z4 hypomethylation due to mutation of DNMT3B (ICF syndrome); and normal or cancer tissues. However, RNAPol-ChIP assays indicated transcription of D4Z4 in a chromosome 4-containing human-mouse somatic cell hybrid. ChIP and RT-PCR showed satellite DNA transcription in some cancers and lymphoblastoid cell lines, although only at a low level. Given the evidence for the involvement of RNAi in satellite DNA heterochromatinization, it is surprising that, at most, a very small fraction of satellite DNA was associated with RNA Pol II. In addition, our results do not support the previously hypothesized disease-linked differential transcription of D4Z4 sequences in short, FSHD-linked arrays.
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PMID:RNAPol-ChIP analysis of transcription from FSHD-linked tandem repeats and satellite DNA. 1723 56

Potassium is an essential element for plant, and high-affinity K+ uptake system plays a crucial role in potassium absorption and transportation. Here we report the isolation and characterization of a HKT1 homolog from C3 halophyte Suaeda salsa (L.) (SsHKT1), particularly under low K+ treatment. The SsHKT1 cDNA was 2033 nucleotides long including 1650 bp ORF for a 550 amino acids peptide and a predicted molecular mass of 63.0 kDa. The deduced amino acid sequence of SsHKT1 was 39-64% identical to other plant HKT-like sequences. A SsHKT1-specific antibody was prepared and reacted with a 63.0 kDa protein from S. salsa plasma membrane. Reverse transcriptase-PCR analysis showed that SsHKT1 was mainly expressed in leaf tissues and to a lesser extent, in root tissues. Amounts of SsHKT1 transcript were developmentally controlled and significantly up-regulated by K+ deprivation and NaCl treatment. The results suggested that SsHKT1 might play an important role in ion homeostasis and salt tolerance of S. salsa.
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PMID:Cloning and expression pattern of SsHKT1 encoding a putative cation transporter from halophyte Suaeda salsa. 1785 52

The open reading frame (ORF3) genes of the parent DR13, attenuated DR13, KPED-9, P-5V, and 12 field samples were cloned and sequenced to further explore the functions of wild- and attenuated-type porcine epidemic diarrhea viruses (PEDVs). Sequencing revealed that wild-type PEDVs ORF3 genes had a single ORF of 675 nucleotides encoding a protein of 224 amino acids with a predicted M (r) of 25.1-25.3 kDa. Attenuated-type PEDVs ORF3 genes had a single ORF of 624 nucleotides encoding a protein of 207 amino acids with a predicted M (r) of 23.4 kDa. The coding region of the ORF3 gene of attenuated-type PEDVs including attenuated DR13, KPED-9, and P-5V had 51 nucleotide deletions that were not found in the ORF3 genes of wild-type PEDVs including CV777, Br1/87, LZC, parent DR13, and 12 field samples. In addition, attenuated-type PEDVs have previously been found to exhibit reduced pathogenicity in pigs. Therefore, 51 nucleotide deletions appear to be meaningful and may be significant for PEDV pathogenicity, because they lead to changes in the predicted amino acid sequences of attenuated-type PEDVs. Reverse transcriptase-polymerase chain reaction (RT-PCR) on the partial ORF3 gene including 51 nucleotide deletions revealed that all PEDVs fell into two types, wild- and attenuated-type PEDVs. Wild-type PEDVs containing parent DR13 and 12 field samples had RT-PCR products of 245 bp in size, while attenuated-type PEDVs containing PEDV vaccine strains (attenuated DR13, KPED-9, P-5V) had products of 194 bp. In addition, all PEDV vaccine strains were used as live virus vaccine, because they previously exhibited a reduced pathogenicity in pigs. Therefore, large deletion region, which is comprise 17 amino acid deletions caused by 51 nucleotide deletions and is seen in all PED live vaccine strains, may be important site for PEDV pathogenicity, and we can use it for differentiation of wild- and attenuated-type PEDVs.
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PMID:Cloning and further sequence analysis of the ORF3 gene of wild- and attenuated-type porcine epidemic diarrhea viruses. 1793 36

The aim of this study was to identify tissues where ovine herpesvirus 2 (OvHV-2) replication occurs in vivo. A reverse-transcriptase PCR targeting the OvHV-2 major capsid protein gene (ORF 25) was developed and the presence of transcripts used as an indicator of virus replication in naturally infected sheep, and cattle and bison with sheep-associated malignant catarrhal fever (SA-MCF). ORF 25 transcripts were detected in 18 of 60 (30%) turbinate, trachea, and lung samples from five sheep experiencing a shedding episode; 12 of the 18 positive samples were turbinates. ORF 25 transcripts were not detected in any other tissue from the shedding sheep (n=55). In contrast, 86 of 102 (84%) samples from clinically affected bovine and bison tissues, including brain, kidney, intestine, and bladder, had ORF 25 transcripts. The data strongly suggest that OvHV-2 replication is localized to the respiratory tract of shedding sheep, predominantly in the turbinate, while it occurs in virtually all tissues of cattle and bison with SA-MCF. These findings represent an important initial step in understanding viral pathogenesis, and in potentially establishing a system for OvHV-2 propagation in vitro.
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PMID:Detection of ovine herpesvirus 2 major capsid gene transcripts as an indicator of virus replication in shedding sheep and clinically affected animals. 1805 5

A 10.3kb linear mitochondrial DNA plasmid designated pFP1 was isolated from Fusarium proliferatum. The DNA sequence of the plasmid consists of 10,336bp with perfect terminal inverted repeats of 400bp. Two major, non-overlapping ORFs were identified on opposite strands, encoding a phage-type RNA polymerase and a family B type DNA polymerase, respectively. One additional minor ORF encoding a putative highly basic protein was also identified. The copy number of pFP1, as determined by RT-PCR, ranged between 1.8 and 3.1 per mtDNA copies depending on the host strain. Real-time PCR analysis of a total of 400 cultures surviving ethidium bromide curing indicated that no plasmid-free strains could be obtained by this treatment. Further single spore selections of the survivors with reduced plasmid content were needed to obtain plasmid-free clones. No phenotypic differences were found between the wild-type strains and their plasmid-free progenies.
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PMID:Characterization of a new mitochondrial plasmid from Fusarium proliferatum. 1826 47

The cAMP response element-binding protein (CREB) is a transcription factor that plays important roles in cellular growth, proliferation and survival. Here, we report that a homologue of CREB transcription factor, Ca-CREB, was identified and functionally characterized in oyster, Crassostrea ariakensis. The full-length cDNA consists of 1397bp with an ORF encoding a 39.3kDa protein. Amino acid sequence analysis revealed that Ca-CREB shares conserved signature motifs with other CREB proteins. Ca-CREB was ubiquitously and constitutively expressed in oyster, and the expression level in hemocytes was higher than that in other tissues. The expression level of Ca-CREB was not modified after RLO stimulation, while tumor necrosis factor-alpha (TNF-alpha) expression was increased obviously, which was revealed by real-time reverse-transcriptase polymerase chain reaction (RT-PCR). Electrophoretic mobility shift assay (EMSA) and Western blotting showed that recombinant CREB proteins specifically bind the consensus CREB binding site, and DNA-binding activity and phosphorylation of Ca-CREB were induced by RLO. These results suggest that Ca-CREB is a CREB homologue and may be involved in immune responses against RLO.
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PMID:Characterization and function of CREB homologue from Crassostrea ariakensis stimulated by rickettsia-like organism. 1860 51

During the outbreak of SARS in 2002/3, a prototype virus was isolated from a patient in Frankfurt/Germany (strain Frankfurt-1). As opposed to all other SARS-Coronavirus strains, Frankfurt-1 has a 45-nucleotide deletion in the transmembrane domain of its ORF 7b protein. When over-expressed in HEK 293 cells, the full-length protein but not the variant with the deletion caused interferon beta induction and cleavage of procaspase 3. To study the role of ORF 7b in the context of virus replication, we cloned a full genome cDNA copy of Frankfurt-1 in a bacterial artificial chromosome downstream of a T7 RNA polymerase promoter. Transfection of capped RNA transcribed from this construct yielded infectious virus that was indistinguishable from the original virus isolate. The presumed Frankfurt-1 ancestor with an intact ORF 7b was reconstructed. In CaCo-2 and HUH7 cells, but not in Vero cells, the variant carrying the ORF 7b deletion had a replicative advantage against the parental virus (4- and 6-fold increase of virus RNA in supernatant, respectively). This effect was neither associated with changes in the induction or secretion of type I interferon, nor with altered induction of apoptosis in cell culture. However, pretreatment of cells with interferon beta caused the deleted virus to replicate to higher titers than the parental strain (3.4-fold in Vero cells, 7.9-fold in CaCo-2 cells). In Syrian Golden Hamsters inoculated intranasally with 10e4 plaque forming units of either virus, mean titers of infectious virus and viral RNA in the lungs after 24 h were increased 23- and 94.8-fold, respectively, with the deleted virus. This difference could explain earlier observations of enhanced virulence of Frankfurt-1 in Hamsters as compared to other SARS-Coronavirus reference strains and identifies the SARS-CoV 7b protein as an attenuating factor with the SARS-Coronavirus genome. Because attenuation was focused on the early phase of infection in-vivo, ORF 7b might have contributed to the delayed accumulation of virus in patients that was suggested to have limited the spread of the SARS epidemic.
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PMID:Reverse genetic characterization of the natural genomic deletion in SARS-Coronavirus strain Frankfurt-1 open reading frame 7b reveals an attenuating function of the 7b protein in-vitro and in-vivo. 1969 90

The locus alx, which encodes a putative transporter, was discovered previously in a screen for genes induced under extreme alkaline conditions. Here we show that the RNA region preceding the alx ORF acts as a pH-responsive element, which, in response to high pH, leads to an increase in alx expression. Under normal growth conditions this RNA region forms a translationally inactive structure, but when exposed to high pH, a translationally active structure is formed to produce Alx. Formation of the active structure occurs while transcription is in progress under alkaline conditions and involves pausing of RNA polymerase at two distinct sites. Alkali increases the longevity of pausing at these sites and thereby interferes with formation of the inactive structure and promotes folding of the active one. The alx locus represents the first example of a pH-responsive riboregulator of gene expression, introducing a novel regulatory mechanism that involves RNA folding dynamics driven by pH.
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PMID:A pH-responsive riboregulator. 1993 54

The development of a reverse transcriptase-polymerase chain reaction (RT-PCR) test for detecting chicken astroviruses (CAstV) is described. Primers, which amplified a fragment of 510 base pairs, were located in conserved regions within the ORF 1b (RNA polymerase) gene. The limit of detection of the test was estimated to be approximately 60 viral copies using a 10-fold dilution series of in vitro transcribed RNA. Positive signals were produced with representative CAstV samples, some of which were not detected by a previously described RT-PCR test for detecting CAstV, but other avian astroviruses including avian nephritis virus and duck hepatitis virus types 2 and 3 tested negative. When applied to gut content samples and swabs from UK and German broiler flocks with growth problems, CAstVs were detected by RT-PCR in 50/52 (96%) samples. CAstVs were detected in between 30% and 72.5% pooled gut content samples from longitudinal surveys of four broiler flocks displaying below-average performances. Whereas all day 0 samples were CAstV-negative, high detection rates were observed when the surveyed birds were aged 4, 5 and 7 days. Based on partial ORF 1b sequences, a phylogenetic analysis of 20 CAstVs indicated the existence of two groups. One group comprised four CAstV isolates, including FP3 and 11672, and two field CAstVs, which shared >94% nucleotide identity. The remaining 14 CAstVs, comprising the first characterized CAstV and 612 isolates and 12 field CAstVs, shared 85% to 99% nucleotide identity and displayed 76% to 79% nucleotide identity with the 11672-like and FP3-like CAstVs.
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PMID:Detection of chicken astrovirus by reverse transcriptase-polymerase chain reaction. 1993 14

The biological impact of transposons on the physiology of the host depends greatly on the frequency and position of integration. Previous studies of Tf1, a long terminal repeat retrotransposon in Schizosaccharomyces pombe, showed that integration occurs at the promoters of RNA polymerase II (Pol II) transcribed genes. To determine whether specific promoters are preferred targets of integration, we sequenced large numbers of insertions using high-throughput pyrosequencing. In four independent experiments we identified a total of 73,125 independent integration events. These data provided strong support for the conclusion that Pol II promoters are the targets of Tf1 integration. The size and number of the integration experiments resulted in reproducible measures of integration for each intergenic region and ORF in the S. pombe genome. The reproducibility of the integration activity from experiment to experiment demonstrates that we have saturated the full set of insertion sites that are actively targeted by Tf1. We found Tf1 integration was highly biased in favor of a specific set of Pol II promoters. The overwhelming majority (76%) of the insertions were distributed in intergenic sequences that contained 31% of the promoters of S. pombe. Interestingly, there was no correlation between the amount of integration at these promoters and their level of transcription. Instead, we found Tf1 had a strong preference for promoters that are induced by conditions of stress. This targeting of stress response genes coupled with the ability of Tf1 to regulate the expression of adjacent genes suggests Tf1 may improve the survival of S. pombe when cells are exposed to environmental stress.
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PMID:High-throughput sequencing of retrotransposon integration provides a saturated profile of target activity in Schizosaccharomyces pombe. 2004 May 83


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