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)

DNA extraction from the rumen of three species of goat (boer goat, Nanjiang yellow goat, Inner Mongolia cashmere goat) was followed by Polymerase Chain Reaction (PCR) amplification of the beta subunit of the RNA polymerase (rpoB) and 16S rDNA genes. PCR products were compared by Denaturing Gradient Gel Electrophoresis (DGGE) to compare the predominant bacterial community structure. The results showed the rpoB DGGE profiles comprised fewer bands than those of 16S rDNA profiles and were easier to analyze. The gene for rpo B is a single copy gene unlike 16S rDNA. So using the rpoB gene offeres a better alternative to the commonly used 16S rRNA gene for microbial community analyses based on DGGE. The bacteria community structure of different goats were similar to each other. The similarities within species were noticeably higher than that between species. Goat species were found to influence the rumen microbe community. Phylogenetic and sequence similarity analyses of the resultant 14 clone sequences in16S rDNA DGGE libraries revealed that 4 clone show similarity over 97% with that of database sequences, while the rest present similarity in a range of 89%-96%, and 13 clone of all were similar to those unidentified rumen bacteria. These results suggest that DGGE followed by clone technique is a practicable protocol to research the complex community of rumen microbe.
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PMID:[Use of rpoB and 16S rDNA genes to analyze rumen bacterial diversity of goat using PCR and DGGE]. 1755 36

Heterochromatin formation involves the nucleation and spreading of structural and epigenetic features along the chromatin fiber. Chromatin barriers and associated proteins counteract the spreading of heterochromatin, thereby restricting it to specific regions of the genome. We have performed gene expression studies and chromatin immunoprecipitation on strains in which native centromere sequences have been mutated to study the mechanism by which a tRNA(Alanine) gene barrier (cen1 tDNA(Ala)) blocks the spread of pericentromeric heterochromatin at the centromere of chromosome 1 (cen1) in the fission yeast, Schizosaccharomyces pombe. Within the centromere, barrier activity is a general property of tDNAs and, unlike previously characterized barriers, requires the association of both transcription factor IIIC and RNA Polymerase III. Although the cen1 tDNA(Ala) gene is actively transcribed, barrier activity is independent of transcriptional orientation. These findings provide experimental evidence for the involvement of a fully assembled RNA polymerase III transcription complex in defining independent structural and functional domains at a eukaryotic centromere.
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PMID:An RNA polymerase III-dependent heterochromatin barrier at fission yeast centromere 1. 1797 62

The DNA lesion 1,N(2)-ethenoguanine (1,N(2)-epsilon G) is formed endogenously as a by-product of lipid peroxidation or by reaction with epoxides that result from the metabolism of the industrial pollutant vinyl chloride, a known human carcinogen. DNA replication past 1,N(2)-epsilon G and site-specific mutagenesis studies on mammalian cells have established the highly mutagenic and genotoxic properties of the damaged base. However, there is as yet no information on the processing of this lesion during transcription. Here, we report the results of transcription past a site-specifically modified 1,N(2)-epsilon G DNA template. This lesion contains an exocyclic ring obstructing the Watson-Crick hydrogen-bonding edge of guanine. Our results show that 1,N(2)-epsilon G acts as a partial block to the bacteriophage T7 RNA polymerase (RNAP), which allows nucleotide incorporation in the growing RNA with the selectivity A>G>(C=-1 deletion)>>U. In contrast, 1,N(2)-epsilon G poses an absolute block to human RNAP II elongation, and nucleotide incorporation opposite the lesion is not observed. Computer modeling studies show that the more open active site of T7 RNAP allows lesion bypass when the 1,N(2)-epsilon G adopts the syn-conformation. This orientation places the exocyclic ring in a collision-free empty pocket of the polymerase, and the observed base incorporation preferences are in agreement with hydrogen-bonding possibilities between the incoming nucleotides and the Hoogsteen edge of the lesion. On the other hand, in the more crowded active site of the human RNAP II, the modeling studies show that both syn- and anti-conformations of the 1,N(2)-epsilon G are sterically impermissible. Polymerase stalling is currently believed to trigger the transcription-coupled nucleotide excision repair machinery. Thus, our data suggest that this repair pathway is likely engaged in the clearance of the 1,N(2)-epsilon G from actively transcribed DNA.
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PMID:Transcription processing at 1,N2-ethenoguanine by human RNA polymerase II and bacteriophage T7 RNA polymerase. 1802 39

B cells produce Ig H chain (IgH) mRNA and protein, primarily of the membrane-bound specific form. Plasma cells produce 20- to 50-fold higher amounts of IgH mRNA, most processed to the secretory specific form; this shift is mediated by substantial changes in RNA processing but only a small increase in IgH transcription rate. We investigated RNA polymerase II (RNAP-II) loading and phosphorylation of its C-terminal domain (CTD) on the IgG2a H chain gene, comparing two mouse cell lines representing B (A20) and plasma cells (AxJ) that express the identical H chain gene whose RNA is processed in different ways. Using chromatin immunoprecipitation and real-time PCR, we detected increased RNAP-II and Ser-2 and Ser-5 phosphorylation of RNAP-II CTD close to the IgH promoter in plasma cells. We detected increased association of several 3' end-processing factors, ELL2 and PC4, at the 5' end of the IgH gene in AxJ as compared with A20 cells. Polymerase progress and factor associations were inhibited by 5,6-dichlorobenzimidazole riboside, a drug that interferes with the addition of the Ser-2 to the CTD of RNAP-II. Taken together, these data indicate a role for CTD phosphorylation and polyadenylation/ELL2/PC4 factor loading on the polymerase in the choice of the secretory poly(A) site for the IgH gene.
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PMID:Increased phosphorylation of the carboxyl-terminal domain of RNA polymerase II and loading of polyadenylation and cotranscriptional factors contribute to regulation of the ig heavy chain mRNA in plasma cells. 1802 12

A novel Eschericha coli expression system directed by bacteriophage T7 RNA Polymerase utilized for overexpression of the cloned gene. The recombinant cell contains the plasmid with a bacteriophage promoter, the T7 promoter, to regulate the expression of the target gene. This promoter is recognized only by T7 RNA polymerase, whose gene has been fused into the host chromosome and is under control of the lacUV5 promoter. Therefore, the target gene on the plasmid can be expressed only in the presence of T7 RNA polymerase, which is induced by isopropyl-beta-D-thiogalactopyranoside (IPTG). The batch cultures were performed to investigate the effect of induction on kinetics of cell growth and foreign protein formation and to determine the optimal induction strategy. It was observed that the specific growth rates of the recombinant cells dramatically decrease after induction, and that there is an optimal induction time for maximizing the accumulated intracellular foreign protein. This optimal induction time varies significantly with inducer concentration. To better understand the optimal behavior, a lumped mechanistic model was constructed to analyze the induced cell growth and foreign protein formation rates.
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PMID:Overexpression of cloned genes using recombinant Escherichia coli regulated by a T7 promoter: I. Batch cultures and kinetic modeling. 1860 Nov 82

The four members of the albumin gene family encode the serum transport proteins albumin, alpha-fetoprotein, alpha-albumin, and vitamin D-binding protein. These genes are transcribed primarily in the liver with each having a different pattern of developmental expression. The tight linkage of these genes, particularly that of albumin, alpha-fetoprotein and alpha-albumin, and their liver-specific expression, has led to the suggestion that these genes share common regulatory elements. To directly examine whether the alpha-fetoprotein enhancer region could regulate the albumin gene family, expression of these genes was monitored in mice in which this region was deleted by homologous recombination. Our data indicate that this enhancer region is required for alpha-fetoprotein and albumin activation early in liver development and alpha-fetoprotein reactivation during liver regeneration, but that albumin, alpha-albumin, and vitamin D-binding protein expression later in hepatic development is not affected by the absence of these enhancers. We also demonstrate that RNA polymerase II loading on the alpha-fetoprotein and albumin promoters is reduced in the absence of this enhancer region, indicating a direct role for these enhancers in the assembly of the RNA Polymerase II complex during liver development.
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PMID:The alpha-fetoprotein enhancer region activates the albumin and alpha-fetoprotein promoters during liver development. 1978 60

There are two recognized poxviruses that are associated with disease in tree squirrels: squirrel fibroma virus (SQFV), Leporipoxvirus, which affects eastern grey squirrels (Sciurus carolinensis) in eastern North America, and squirrelpox virus (SQPV), a member of a newly identified poxvirus genus, which affects European red squirrels (Sciurus vulgaris) in the United Kingdom. In August 2008, a cutaneous poxvirus-associated disease was identified in a North American red squirrel (Tamiasciurus hudsonicus) from the Yukon Territory, Canada. The gross and microscopic appearance of the skin lesions was more consistent with SQPV than SQFV, and electron microscopy revealed poxvirions only within epithelial cells. Polymerase chain reaction (PCR) was used to identify poxvirus core protein encoding DNA in skin samples, and phylogenetic analysis showed that the inferred amino acid sequence was distinct from all other poxvirus species for which the core protein gene has been sequenced, including those of the genus Leporipoxvirus. Although the core protein sequence of SQPV was not available, comparison of the constructed phylogenetic tree to other published trees, based on major outer envelope proteins, revealed that the identified sequence occupies a position similar to SQPV in terms of its relationship to other poxviruses. However, PCR primers designed to amplify gene sequences encoding the SQPV major envelope protein and RNA polymerase did not amplify any sequences from infected tissues. These findings suggest that the virus present in this squirrel is a novel poxvirus of North American red squirrels. To our knowledge, this is the first case of poxvirus infection in Canadian squirrels outside of Ontario.
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PMID:Poxvirus infection in an American red squirrel (Tamiasciurus hudsonicus) from northwestern Canada. 1990 87

Triplex-forming oligonucleotides (TFOs) are attractive tools to control gene expression at the transcriptional level. This anti-gene approach has proven to be successful in various experimental settings. However, the mechanisms leading to transcriptional repression in cells have not been fully investigated yet. Here, we examined the consequence of triplex DNA formation on the binding of transcriptional activators, co-activators and RNA Polymerase II to the ets2 gene promoter using chromatin immunoprecipitation assays. The triplex target sequence was located approximately 40-bp upstream of the transcription start site (TSS) and overlapped an Sp1 binding site relevant for ets2 transcription. We found that the ets2-TFO prevented binding of Sp1, TAF(II)130 and TAF(II)250 to the ets2 promoter, while binding of RNA polymerase II and TBP were not affected. The effects were both sequence and target specific, since the TFO had no effect on the c-myc promoter and a mutated ets2 promoter construct. Thus, triplex DNA formation near a TSS leads to formation of a non-functional pre-initiation complex (PIC) by blocking binding of transcriptional activators and co-activator molecules. This is the first direct demonstration of interference with PIC assembly at the TSS by oligonucleotide-triplex DNA formation in cells.
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PMID:Mechanisms of triplex DNA-mediated inhibition of transcription initiation in cells. 2004 41

Capping of mRNAs is strictly coupled to RNA polymerase II transcription and there is evidence, mainly from metazoans, that other steps in pre-mRNA processing show a similar linkage. In trypanosomes, however, the mRNA cap is supplied by a trans spliced leader sequence. Thus pre-mRNAs transcribed by RNA Polymerase I are capped by trans splicing, and translation-competent transgenic mRNAs can be produced by RNA Polymerase I and T7 RNA polymerase so long as the primary transcript has a splice acceptor signal. We quantified the efficiency of processing of trypanosome pre-mRNAs produced from a plasmid integrated either at the tubulin locus, or in an rRNA spacer, and transcribed by RNA polymerase II, RNA polymerase I or T7 RNA polymerase. The processing efficiencies were similar for primary transcripts from the tubulin locus, produced by RNA polymerase II, and for RNA from an rRNA spacer, transcribed by RNA polymerase I. Primary transcripts produced by T7 RNA polymerase from the tubulin locus were processed almost as well. There was therefore no evidence for recruitment of the 3'-splicing apparatus by the RNA polymerase. Abundant transcripts transcribed from the rRNA locus by T7 RNA polymerase were somewhat less efficiently processed.
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PMID:Processing of a phosphoglycerate kinase reporter mRNA in Trypanosoma brucei is not coupled to transcription by RNA polymerase II. 2036 63

In this article, we report the resonance assignment of CTD-interacting domain (CID) of pre-mRNA down-regulation (Nrd)1 bound to Ser5-phosphorylated CTD (pSer5) of RNA Polymerase II. The presented assignment of backbone and side-chain resonances of the Nrd1 CID proton, carbon and nitrogen nuclei will allow studies of the structure and interaction of CID with carboxy-terminal domain (CTD) of the RNA polymerase II.
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PMID:1H, 13C, and 15N resonance assignments for the CTD-interacting domain of Nrd1 bound to Ser5-phosphorylated CTD of RNA polymerase II. 2135 Sep 22


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