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)

Temperature-sensitive "leaky" mutants of phage MS2 having white dense ring around negative colonies are described. As these mutants are used for quantitative genetic studies, the white ring presents interest. Typical mutant 40 is used as a model for investigation. Light microscopy has shown, that cells from white ring zone have spore-like inclusions, which determine the characteristic structure of surrounding mutant negative colonies. Cytochemical reactions for the presence of glicogen, lipids, volutin, nuclear material and spores were negative. Electrone microscopy of negatively stained samples and ultrathin sections has revealed that cells from white ring zone, unlike phage-infected wild type cells, have two types of electron dense inclusions: 1) crystalline structures formed with great number of closely packed mature phage particles, and 2) large amorphic bodies. Electrone microscope-cytochemical data showed that inclusions remain intact under treatment of ultrathin sections of white zone ring with DNase and perchloric acid, while nuclear material was completely destroyed. Amorphic bodied were completely destructed after the treatment with RNase, while nuclear material and crystalline phage aggregated remained unchanged. Therefore, amorphic bodies consist of RNA, which has not been used to form virions. Single cycle of the development of mutant 40 at 37 degrees and 43 degrees C and under the temperature of incubation 37 degrees leads to 43 degrees C and 43 degrees leads to 37 degrees C in the course of intracellular reproduction is investigated. Influence of the phage on growth on infected culture is studied. The data obtained draw to a conclusion that the impaired function belongs to cystron protein of the phage membrane. As certain mutations in this cystrone of RNA-containing phage result in the depression of cystrone RNA polymerase, it is supposed that the formation of RNA containing bodies in infected cells, determining the formation of white rings in NA, together with cristalline aggregates of cells, is a result of mutation damage of cystrone protein of the phage MS2 membrane.
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PMID:[Effect of mutagens on RNA-containing phages and its infectious RNA. VII. Genetic nature of morphologic mutants of RNA-containing phage MS2]. 99 65

DNA-dependent RNA polymerase was isolated from rat spleen cell nuclei and was identified as A and B RNA polymerases by data on DEAE- and P-cellulose ionic exchange chromatography and on concentration dependency on bivalent ions and (NH4)2SO4. Two forms of the enzyme differed from each other in the activity in RNA synthesizing system, and their activity was completely inhibited by actinomycin, DNase and RNase.
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PMID:[DNA-dependent RNA polymerase from the nuclei of the spleen of white rats]. 113 3

Ribonucleic acid (RNA)-dependent RNA polymerase activity was demonstrated in the microsomal and ribosomal fraction from the spleen cells of immunized mice. The enzyme activity was solubilized by Triton X-100 from the fraction and partially purified by Biogel A 1.5 m column chromatography. The RNA-dependent RNA polymerase activity was eluted in a single peak from the column. High activity was demonstrated with an RNA polymerase activity was eluted in a single peak from the column. High activity was demonstrated with an RAN preparation (iotaRNA) as template made from the spleens of immunized mice but very low activity was found with an RNA preparation made from the spleens of normal mice. Incorporation of 3H-UTP markedly decreased in the presence of RNase but not in the presence of DNase. DNA preparations made from the spleens of immunized mice were inactive as template for this enzyme. The iotaRNA preparation was fractionated by sucrose density gradient centrifugation. A fraction corresponding to 12-13 S was most active as a template. It was followed by a fraction corresponding to 6-7 S. Sucrose gradient analysis of the 3H-UTP-labeled product was attempted. Some properties of this enzyme are described.
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PMID:Ribonucleic acid-dependent ribonucleic acid polymerase in the immune response. 123 May 9

DNA-dependent RNA polymerase from Escherchia coli was used to transcribe chromatin from human leukocytes and purified human DNA. RNA was labeled at the 5' terminus with either [gamma-32P]ATP or [gamma-32P]GTP and internally with [3H]UTP. Determination of the average chain length of the RNA molecules by the ratio of moles of 3H-labeled nucleotide incorporated to moles 32P-labeled nucleotide incorporated showed that the size of the transcript of purified DNA was about 2 1/2 times greater than those from chromatin. The percentage of chains initiated with ATP and GTP was observed to vary with the template, the ATP to GTP ratio being greater on chromatin. The kinetics of 3H and 32P hybridization of transcripts of purified DNA showed hybridization primarily to nonrepetitive sequences. Transcripts from the chromatin templates when hybridized to DNA showed a larger proportion of RNase resistance of the 32P-termini at low Cot's.
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PMID:Template restriction in human chromatin. 126 Aug 58

We have cloned and determined the nucleotide (nt) sequence of a 6.5-kb genomic DNA fragment containing the rat MyoD gene (encoding a muscle regulatory factor, MyoD). Mouse fibroblasts transfected with this DNA display a high degree of conversion to a muscle phenotype, suggesting that this genomic clone contains sufficient sequence information to allow the production of the rat MyoD protein in these cells. The 6.5-kb genomic fragment contains the complete coding region of MyoD, distributed over three exons, plus 2.3 kb of 5'-noncoding sequence and 1.4 kb of 3'-noncoding sequence. Based on RNase protection assays, the major transcription start point of MyoD is located 210 nt 5' to a methionine start codon and 26 nt 3' to a TAAATA motif which bears similarity to a consensus recognition sequence (TATA) utilized by eukaryotic RNA polymerase II transcription complexes. The high degree of identity between the amino acid sequence of rat MyoD and the MyoD proteins isolated from other vertebrates indicates that this muscle regulatory protein has been evolutionarily conserved.
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PMID:Isolation and structural analysis of the rat MyoD gene. 132 78

Insulin-like growth factors I and II (IGF I and II) are polypeptides with both growth-promoting and insulin-like metabolic effects. Immunoreactive IGF I is present in the retina and both IGF I and II are present in vitreal fluid. The type I and type II IGF receptors are also localized within the neural retina. The presence of IGFs and IGF receptors within the eye suggests a possible growth-promoting effect of IGFs on ocular tissues. IGF may enter the eye from the blood or, alternatively, arise from an ocular cell type which synthesizes and secretes IGF. IGF I and II mRNA synthesis in scleral cells and IGF I synthesis in rat retina suggests endogenous IGF production in the eye. We hypothesized that IGFs and IGF receptors are synthesized by one ocular cell type, the retinal pigment-epithelium (RPE). As a first step in studying IGF production by the RPE, we analyzed expression of the IGF and IGF receptor genes by cultured human RPE cells. Using Northern analysis, RNase protection and reverse-transcriptase polymerase chain reaction (RT-PCR), we found that cultured RPE cells synthesize mRNA for IGF I and the type I and type II IGF receptors.
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PMID:Gene expression of the insulin-like growth factors and their receptors in cultured human retinal pigment epithelial cells. 137 66

U14 small nuclear RNA (snRNA) is an evolutionarily conserved RNA species that plays a role in rRNA processing. The conserved ability of fungal, amphibian and mammalian U14 snRNAs to hybridize with both homologous and heterologous eukaryotic 18S rRNAs indicates a potential role for this intermolecular RNA/RNA interaction in U14 snRNA function. To understand better the possible role of this intermolecular base-pairing in rRNA processing, we have defined those nucleotide sequences in mouse U14 snRNA and 18S rRNA responsible for the observed in vitro hybridization. We have constructed, using synthetic DNA oligonucleotides, a U14 snRNA gene which has been positioned behind a T7 RNA polymerase promoter site and then inserted into a plasmid. The presence of natural or engineered restriction endonuclease sites within this construct has permitted the in vitro transcription of full-length mouse U14 snRNA transcripts (an 87-nucleotide mouse U14 snRNA minus 5' or 3' leader sequences) or 3' terminally truncated U14 snRNA fragments. Hybridization of full-length or truncated fragments of U14 snRNA to mouse 18S rRNA demonstrated the utilization of a previously proposed 18S rRNA complementary sequence located near the 3' end of mouse U14 snRNA (nucleotides 65-78) for intermolecular hybridization. Conversely, RNase-T1-generated fragments of 18S rRNA capable of hybrid-selection by U14 snRNA have been isolated and sequenced. A nested set of hybrid-selected 18S rRNA fragments define a mouse 18S rRNA sequence (nucleotides 459-472) which exhibits perfect complementarity to the defined U14 snRNA sequence 65-78. Primer-extension/chain-termination mapping of mouse U14-snRNA.18S-rRNA hybrids has confirmed the formation of the proposed hybrid structure. A second set of observed complementary sequences in mouse U14 snRNA (nucleotides 25-38) and mouse 18S rRNA (nucleotides 82-95) are not used for the in vitro hybridization of these two RNAs. Presumably the involvement of this second 18S-rRNA-complementary sequence in the secondary/tertiary folding of mouse U14 snRNA prevents its base-pairing with 18S rRNA. However, the strong evolutionary conservation of both U14-snRNA.18S-rRNA hybrid structures and their juxtapositioning within the folded secondary structure of 18S rRNAs argues for a biological role for each in U14 snRNA function.
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PMID:Determination of the nucleotide sequences in mouse U14 small nuclear RNA and 18S ribosomal RNA responsible for in vitro intermolecular base-pairing. 137 13

RNase MRP is a site-specific ribonucleoprotein endoribonuclease that cleaves RNA sequence complementary to mammalian mitochondrial origins of replication in a manner consistent with a role in primer RNA metabolism. The same activity in the yeast Saccharomyces cerevisiae has recently been identified; it cleaves an RNA substrate complementary to a yeast mitochondrial origin of replication at an exact site of linkage of RNA to DNA. We have purified this yeast enzyme further and detect a single, novel RNA of 340 nucleotides associated with the enzymatic activity. The single-copy nuclear gene for this RNA was sequenced and mapped to the right arm of chromosome XIV. The identity of the clone, as encoding the RNA copurifying with enzymatic activity, was confirmed by a match to the directly determined sequence of the RNA. The gene sequence also identified a 340-nucleotide RNA in total yeast RNA and in purified RNase MRP enzyme preparations. Inspection of the sequence of the yeast RNA revealed homologies to the RNA component of mouse RNase MRP, 49% overall with specific regions of much greater similarity. The flanking regions of the gene showed characteristics of an RNA polymerase II transcription unit, including a TATAAA box and a 7/8 match to the yeast cell cycle box UAS. The RNase MRP RNA gene was deleted by insertional replacement and found to be essential for cellular viability, indicating a critical nuclear role for RNase MRP.
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PMID:Yeast site-specific ribonucleoprotein endoribonuclease MRP contains an RNA component homologous to mammalian RNase MRP RNA and essential for cell viability. 139 74

We have developed a modified RNase protection assay in which the antisense RNA probe is prepared from a PCR-amplified DNA template rather than from a linearized plasmid DNA template. In this assay, an RNA polymerase promoter sequence is attached to the 5' end of the antisense PCR primer. Using this modified antisense primer in conjunction with the paired sense primer, PCR amplification generates a linear DNA template that includes an RNA polymerase promoter sequence. Transcription in vitro initiated by the incorporated promoter in the presence of RNA polymerase and ribonucleotide triphosphates produces a radiolabeled run-off antisense RNA transcript, which can then be used as probe for RNase protection analysis. Probes generated by this method obviate the need to subclone DNA sequences into transcription vectors for synthesis of antisense transcripts. Due to the simplicity of its design and the lack of need for subcloning, this strategy offers greater flexibility than conventional methods for the production of single-stranded RNA probes, and thus facilitates the implementation of the ribonuclease protection assay.
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PMID:Application of the polymerase chain reaction to the ribonuclease protection assay. 147 48

This report describes a novel assay involving the polymerase chain reaction (PCR) and RNase protection for the rapid and sensitive detection of malignant lymphoid cells by nucleotide sequences within their individual rearranged gamma T-cell receptor (TCRG) genes. In this assay, clonal rearrangements are amplified from the DNA of diagnostic tumor specimens using a consensus V segment primer and a consensus J segment primer to which the promoter for T7 RNA polymerase has been appended. The PCR product from this amplification is transcribed into a radiolabeled RNA probe. Test RNA transcribed from the opposite DNA strand is synthesized by similar methods from TCRG genes of a subsequent biopsy specimen. The test RNA is hybridized with the probe, and mismatched nucleotide sequences in the RNA hybrids are digested by RNase A. Detection of fully protected probe by means of polyacrylamide gel electrophoresis and autoradiography indicates the presence of malignant cells in the test specimen. Dilution experiments with DNA of cell lines from acute lymphoblastic leukemias (ALLs) show that detection of one tumor cell among 10(5) normal bone marrow cells is usually possible. Residual disease was also successfully detected in several cases of ALL during clinical remission, including detection in one case at the 10(-5) level. The procedure described here may provide a simplified and rapid method for the sensitive diagnosis and monitoring of lymphoid malignancies. This procedure should be applicable to most antigen receptor genes, and unlike most comparable methods, requires neither analysis of nucleotide sequence nor synthesis of tumor-specific oligonucleotide probes or primers.
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PMID:Sensitive detection of clonal antigen receptor gene rearrangements for the diagnosis and monitoring of lymphoid neoplasms by a polymerase chain reaction-mediated ribonuclease protection assay. 165 9


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