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

---

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

The synthesis, isolation, and characterization of a new photo-cross-linking uridine 5'-triphosphate analogue are described. This nucleotide analogue, 5-[(4-azidophenacyl)thio]uridine 5'-triphosphate (5-APAS-UTP), contains an aryl azide group approximately 10 A from the uridine ring. The azide is photoactivated by irradiation at 300 nm, resulting in covalent attachment of the nucleotide to adjacent molecules. The nucleotide can be desulfurated with Raney nickel to cause molecular cleavage between the base and the aryl azide. Desulfuration yields uridine 5'-triphosphate and p-azidoacetophenone. If the analogue is cross-linked to another molecule, desulfuration leaves the analogue's acetophenone group attached to that molecule. This effectively leaves behind a molecular tag on molecules that interact with the uridine analogue either as monomeric nucleotide or as part of an RNA molecule. This nucleotide analogue can be incorporated into internal positions in RNA by transcription in vitro with Escherichia coli RNA polymerase. It can therefore be used to examine interactions between RNA and other molecules (e.g., proteins or nucleic acids). Because the sulfur atom can be selectively removed, the covalent bonds formed between analogue-containing RNA and other molecules can be cleaved, when desired, to facilitate identification of the cross-linked molecules and RNA nucleotides in the cross-linked complex.
...
PMID:Synthesis and characterization of 5-[(4-Azidophenacyl)thio]uridine 5'-triphosphate, a cleavable photo-cross-linking nucleotide analogue. 247 76

Mouse oocytes progress through early meiotic prophase during fetal life and reach the diplotene stage by birth. During prepubertal and reproductive life, oocytes are continuously selected to grow from the pool of small primordial oocytes. Growing oocytes reach full size in 2 weeks, and full-grown oocytes are present in rapidly enlarging follicles for about 5 days before meiotic maturation and ovulation. RNA synthesis during early meiotic prophase, as estimated from [3H]uridine incorporation followed by autoradiography and from electron microscopic analysis of nuclear components, proceeds at a moderate rate throughout except for a brief period in early pachytene when synthesis is low or absent. RNA synthesis continues in primordial oocytes at a moderate rate. Incorporation studies, electron microscopic analyses, and particularly measurements of ongoing RNA polymerase activity (completion of initiated chains as analysed in tissue sections) indicate a distinctly increased rate of synthesis during oocyte growth over that of primordial oocytes, followed by a decline in full-grown oocytes. During growth, this rate increases severalfold. The absolute rate of synthesis of heterogeneous nuclear RNA (using rRNA as a standard) during mid-growth is very rapid, but nevertheless still much lower than that in typical lampbrush chromosomes. Most of the hnRNA turns over with a half-life of about 20 min, as is typical in somatic cells. Newly synthesized mRNA-like RNA enters the cytoplasm at about one-half the rate of rRNA, and about one-third of the ribosomes and one-fourth of the mRNA appear in polysomes. In full-grown oocytes, the rate of synthesis falls distinctly, but a significant level of synthesis continues until it essentially ceases at breakdown of the germinal vesicle. During meiotic prophase, chromosomes are most compact at pachytene and unfold lateral projections as RNA synthesis increases in late pachytene-early diplotene. In primordial oocytes, the diplotene state of chromosomes is obvious in most mammals, but in rodents the chromosomes are more evenly dispersed and are said to be in a dictyate state, although they are still presumably in a diplotene configuration. The chromosome core, which is present in leptotene through early diplotene stages, apparently disappears in the dictyate stage.(ABSTRACT TRUNCATED AT 400 WORDS)
...
PMID:Gene expression during oogenesis and oocyte development in mammals. 248 71

Only three forms of Kunjin virus-specified RNA were isolated from cytoplasm early after the latent period (about 15 hr) viz., 44 S genomic-sized single-stranded RNA, 20 S double-stranded "replicative form" (RF), and 20-28 S partially ribonuclease-resistant (about 70%) "replicative intermediate" (RI). The RF and RI were resolved by electrophoresis in aqueous-agarose gel only following LiCl fractionation. The RI did not enter urea-polyacrylamide gels. After denaturation of untreated or RNase-treated RI and RF, only 44 S RNA was present in electropherograms. RNA polymerase activity at 8 hr postinfection was detected by in vitro assays of cytoplasmic extracts and reached a maximum at 24 hr, the only major labeled product being RF; a trace amount of free 44 S RNA was also produced. These results, and the kinetics of incorporation of [3H]uridine into RI, RF, and 44 S RNA in pulse and pulse-chase experiments, formed the basis of a model in which flavivirus RF functions as a recycling template for semiconservative and (mainly) asymmetric replication, on which only one nascent strand is synthesized per cycle.
...
PMID:Replication strategy of Kunjin virus: evidence for recycling role of replicative form RNA as template in semiconservative and asymmetric replication. 257 39

Antibody molecules directed against RNA polymerase I, the enzyme responsible for rRNA synthesis, were introduced into rat hepatoma cells by red cell-mediated microinjection. Access of the antibodies to the nucleolus, the site of rRNA synthesis, was facilitated by microinjecting mitotic cells. Using indirect immunofluorescence, anti-RNA polymerase I immunoglobulins, but not control immunoglobulins, were found localized in the nucleoli of microinjected cells. To assess whether intracellular antibodies could alter RNA synthesis, cultures were labeled with [3H] uridine at various times after microinjection. Reduction in RNA synthesis, relative to cells microinjected with non-immune immunoglobulins, was observed within three hours. These results demonstrate that antibodies introduced into the cytoplasm of mitotic cells via red cell-mediated microinjection have free access to nuclear components and that they remain functional within the nuclei of living cells.
...
PMID:Reduction in RNA synthesis following red cell-mediated microinjection of antibodies to RNA polymerase I. 258 11

The purpose of this study was to examine effects of aphidicolin and alpha-amanitin on visualization of acridine orange (AO) binding to DNA in rat astrocytoma C6 cells and to discuss briefly the significance of AO chromatin interaction products. Aphidicolin inhibited DNA synthesis but percentage of AO positive cells was approximately 60% of that of the untreated control cells. alpha-Amanitin caused a slight inhibition of RNA synthesis and 3H-uridine incorporation in the treated cells was about 64% of that of the untreated cells, whereas a distinct decrease of the number of AO positive cell nuclei was observed. The results suggest that activity of RNA polymerase II and mRNA synthesis is mainly concerned in visualization of AO chromatin interaction products.
...
PMID:Effects of aphidicolin and alpha-amanitin on visualization of acridine orange binding to DNA in rat astrocytoma C6 cells. 258 4

In studies of the effects of changes in mRNA structure and sequence on the initiation of protein synthesis, we used a generally applicable approach to transcribe reconstructed genes for duck alpha A globin by the bacteriophage SP6 RNA polymerase promoter in a pGEM-2 plasmid vector. The genes were reconstructed such that the first nucleotide to be transcribed, the 5' adenosine, was placed directly adjacent to the SP6 promoter sequence. The 3' ends of the genes were constructed such that cleavage with Ssp 1 endonuclease yielded a template that directed the synthesis of mRNA terminating in a poly A tail containing 56 adenosines and a single 3' uridine. Special conditions using a Mn++ buffer were developed to enable the SP6 RNA polymerase to initiate at the 5' adenosine and synthesize the A-start transcription product. The mRNA could be capped and was subsequently used as an effective template for in vitro translation and synthesis of duck alpha A globin.
...
PMID:Construction of mRNA genes for the synthesis and translation of duck alpha globin mRNA. 263 91

The cytotoxicity of 5-fluorouridine (FUrd) results from actions directed at the synthesis of both DNA and RNA. The role of mRNA as a target for FUrd was investigated by selectively decreasing the incorporation of FUrd into RNA polymerase II transcripts of K-562 erythroleukemia cells, which was accomplished by the addition of alpha-amanitin to cultures of K-562 cells permeabilized with lysolecithin. In these cells alpha-amanitin at concentrations of 1-5 micrograms/ml inhibited the incorporation of [3H]-uridine into polyadenylated RNA by up to 45% and decreased the steady-state levels of two specific mRNAs but had no effect on poly A- RNA synthesis. alpha-Amanitin decreased the incorporation of FUrd into poly A+ RNA by up to 60%. The decrease in FUrd incorporation produced by alpha-amanitin was accompanied by an antagonism of the growth inhibitory effects of the fluorinated pyrimidine nucleoside by the mycotoxin, as measured by both growth in suspension culture and colony formation in 0.12% agar. Antagonism between these agents increased as the concentration of alpha-amanitin was elevated; furthermore, it was sequence-dependent, occurring only when alpha-amanitin preceded FUrd. These findings provide evidence that the actions of FUrd directed against mRNA are antagonized when FUrd incorporation into mRNA transcripts is decreased and that the effects of FUrd on mRNA produce cytotoxic consequences.
...
PMID:RNA polymerase II transcripts as targets for 5-fluorouridine cytotoxicity: antagonism of 5-fluorouridine actions by alpha-amanitin. 273 15

Yeast tRNA ligase, from Saccharomyces cerevisiae, is one of the protein components that is involved in the splicing reaction of intron-containing yeast precursor tRNAs. It is an unusual protein because it has three distinct catalytic activities. It functions as a polynucleotide kinase, as a cyclic phosphodiesterase, and as an RNA ligase. We have studied the binding interactions between ligase and precursor tRNAs containing two photoreactive uridine analogues, 4-thiouridine and 5-bromouridine. When irradiated with long ultraviolet light, RNA containing these analogues can form specific covalent bonds with associated proteins. In this paper, we show that 4-thiouridine triphosphate and 5-bromouridine triphosphate were readily incorporated into a precursor tRNA(Phe) that was synthesized, in vitro, with bacteriophage T7 RNA polymerase. The analogue-containing precursor tRNAs were authentic substrates for the two splicing enzymes that were tested (endonuclease and ligase), and they formed specific covalent bonds with ligase when they were irradiated with long-wavelength ultraviolet light. We have determined the position of three major cross-links and one minor cross-link on precursor tRNA(Phe) that were located within the intron and near the 3' splice site. On the basis of these data, we present a model for the in vivo splicing reaction of yeast precursor tRNAs.
...
PMID:Binding interactions between yeast tRNA ligase and a precursor transfer ribonucleic acid containing two photoreactive uridine analogues. 285 71

Ribavirin 5'-sulfamate, a nucleotide analog, inhibited Semliki Forest virus cytopathology by 50% at 10 microM, whereas ribavirin was inactive at less than or equal to 1 mM. Actinomycin D did not reverse (antagonize) the effect of ribavirin 5'-sulfamate against the virus. The compound inhibited amino acid incorporation into macromolecules of uninfected cells but had no appreciable effect on uridine incorporation. Infected cells treated with actinomycin D and nucleotide analog were inhibited in amino acid and uridine incorporation. The compound blocked the formation of the viral RNA polymerase protein in cells, which could account for the inhibited synthesis of new viral RNA. By electrophoresis, inhibition of the synthesis of viral proteins was more pronounced than the inhibition of cellular polypeptides. The analog inhibited the translation of mRNA to protein. Most animals treated intraperitoneally for 7 days with ribavirin 5'-sulfamate at 20 and 40 mg/kg/day starting 2 h before intraperitoneal Semliki Forest virus inoculation survived the otherwise lethal infection.
...
PMID:Antiviral activity and mode of action of ribavirin 5'-sulfamate against Semliki Forest virus. 285 56

The splicing of a procaryotic precursor RNA transcribed from the T4 phage thymidylate synthase (td) gene with SP6 RNA polymerase was investigated in vitro. The intron excision-cyclization reaction increased progressively to 60 degrees C. Exon ligation, though barely detectable at the lower temperatures, was greatly enhanced at 60 degrees C. Both reactions required Mg2+. The addition of guanosine to the 5' end of an intron-exon II intermediate via a 3',5'-phosphodiester bond was essential for the ligation of exon I to exon II. The added guanosine and the first intron-encoded uridine are subsequently lost as a dinucleotide from the 5' end during cyclization of the linear form of the excised intron RNA. Exon ligation is intramolecular and occurs more readily in the nascent RNA molecule (cotranscriptionally) than in the finished transcript (posttranscriptionally). These data and the identification of various structural elements (P, Q, R, S, E, E') in the td intron that are found typically in eucaryotic class I introns firmly establish the td intron as the first example of class I intron of procaryotic origin.
...
PMID:Mechanism and requirements of in vitro RNA splicing of the primary transcript from the T4 bacteriophage thymidylate synthase gene. 303 75


<< Previous 1 2 3 4 5 6 7 8 9 10 Next >>

---