Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.1 (RNase)
 16,360 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

In all retrovirus systems studied, the leader region of the RNA contains a cis-acting sequence called psi that is required for packaging the viral RNA genome. Since the pol and env genes are dispensable for formation of RNA-containing particles, the gag gene product must have an RNA binding domain(s) capable of recognizing psi. To gain information about which portion(s) of Gag is required for RNA packaging in the avian sarcoma and leukemia virus system, we utilized a series of gag deletion mutants that retain the ability to assemble virus-like particles. COS cells were cotransfected with these mutant DNAs plus a tester DNA containing psi, and incorporation of RNA into particles were measured by RNase protection. The efficiency of packaging was determined by normalization of the amount of psi+ RNA to the amount of Gag protein released in virus-like particles. Specificity of packaging was determined by comparisons of psi+ and psi- RNA in particles and in cells. The results indicate that much of the MA domain, much of the p10 domain, half of the CA domain, and the entire PR domain of Gag are unnecessary for efficient packaging. In addition, none of these deleted regions is needed for specific selection of the psi RNA. Deletions within the NC domain, as expected, reduce or eliminate both the efficiency and the specificity of packaging. Among mutants that retain the ability to package, a deletion within the CA domain (which includes the major homology region) is the least efficient. We also examined particles of the well-known packaging mutant SE21Q1b. The data suggest that the random RNA packaging behavior of this mutant is not due to a specific defect but rather is the result of the cumulative effect of many point mutations throughout the gag gene.
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PMID:Efficiency and selectivity of RNA packaging by Rous sarcoma virus Gag deletion mutants. 805 73

Previous studies from this laboratory have demonstrated that the enhancer 1 binding factor (E1BF), a Ku-related protein, purified from the serum-enriched cells functions as a positive factor in an RNA polymerase (pol I) transcription system. We have now shown that E1BF purified from the serum-deprived cells (E1BFs) can inhibit rDNA transcription completely in a fractionated extract from the cells grown in serum-enriched medium. The suppression of transcription was overcome by the addition of control E1BF (E1BFc). Immunoprecipitation of purified E1BFs by the anti-Ku monoclonal antibody and addition of the supernatant to the transcription reaction mixture prevented the inhibition significantly, whereas immunoprecipitation with the control mouse IgG did not restore the transcription. The transcriptional repressor activity associated with the final DNA affinity column fractions copurified with E1BF. Neither the amount of E1BF nor its promoter binding activity was altered following serum depletion. E1BFs selectively inhibited the initiation of rDNA transcription. The inhibitory activity of E1BFs was not due to a nonspecific RNase activity. These data suggest that E1BF is post-translationally modified following serum starvation of cells, and that the repressor activity of E1BFs is largely responsible for the down-regulation of pol I transcription in serum-deprived cells.
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PMID:Enhancer 1 binding factor (E1BF), a Ku-related protein, is a growth-regulated RNA polymerase I transcription factor: association of a repressor activity with purified E1BF from serum-deprived cells. 809 Jul 77

RNA chain elongation by RNA polymerase is a dynamic process. Techniques that allow the isolation of active elongation complexes have enabled investigators to describe individual steps in the polymerization of RNA chains. This article will describe recent studies of elongation by RNA polymerase II (pol II). At least four types of blockage to chain elongation can be overcome by elongation factor SII: (a) naturally occurring "arrest" sequences, (b) DNA-bound protein, (c) drugs bound in the DNA minor groove, and (d) chain-terminating substrates incorporated into the RNA chain. SII binds to RNA polymerase II and stimulates a ribonuclease activity that shortens nascent transcripts from their 3' ends. This RNA cleavage is required for chain elongation from some template positions. As a result, the pol II elongation complex can repeatedly shorten and reextend the nascent RNA chain in a process we refer to as cleavage-resynthesis. Hence, assembly of large RNAs does not necessarily proceed in a direct manner. The ability to shorten and reextend nascent RNAs means that a transcription impediment through which only half the enzyme molecules can proceed per encounter, can be overcome by 99% of the molecules after six iterations of cleavage-resynthesis. Surprisingly, the boundaries of the elongation complex do not move upstream after RNA cleavage. The physico-chemical alterations in the elongation complex that accompany RNA cleavage and permit renewed chain elongation are not yet understood.
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PMID:Transcription elongation by RNA polymerase II: mechanism of SII activation. 831 68

Several sites of interaction with viral proteins have been mapped in the plus strand of the Saccharomyces cerevisiae double-stranded RNA virus, ScV. These include a site necessary and sufficient for viral particle binding to plus strands (VBS) and a site necessary and sufficient for interference with replication of viral plus strands (INS). We show that the INS and VBS are identical and that they are necessary and sufficient for packaging. One of the viral RNAs has two adjacent VBS, which have additive INS activity. The second VBS has similar affinity for viral particles as the first but its complex with particles exhibits faster dissociation. Binding to the two sites in the viral RNA is independent but equivalent. This may mean two recognition sites per particle. This is what would be expected for two molecules of the protein thought to be responsible for sequence specific recognition, the cap-pol fusion polypeptide, per particle. Comparison of the secondary structures of several binding sites, as determined by site-directed mutagenesis and by ribonuclease mapping of single- and double-stranded regions, reveals a requirement for a specific loop sequence and two stem regions, in which there is no sequence specificity. Contrary to what was described in previous work, a bulge loop, but not a "bulged" A residue is necessary for binding of the second VBS. A minimal region of 30 bases is enough to cause both packaging and replication.
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PMID:RNA structural requirements for RNA binding, replication, and packaging in the yeast double-stranded RNA virus. 833 25

RNA synthesis can be detected in nuclei using modified RNA precursors (Br-UTP) introduced in permeabilized cells. Surprisingly, RNA pol I transcripts are detected only after inhibition of RNA pol II or salt enhancement of RNA pol I activity. By modifying a previously reported protocol, we found that RNA pol I transcripts can be detected selectively or simultaneously with RNA pol II transcripts without any drug treatment. Removing glycerol from the permeabilization and transcription buffers and improving the permeabilization using Triton X-100 revealed RNA pol I transcription in two cell lines (mammalian and Xenopus) and in isolated mouse oocytes. The transcripts were most probably rRNA because they were detected in the nucleoli, digested by RNase, sensitive to actinomycin D, and resistant to alpha-amanitin. We found by microinjection of the Br-UTP precursors in living cells that low ionic strength allows the detection of RNA pol I transcription. Electron microscopy of mouse oocytes showed that the "looseness" of the nucleolar organization is associated with the detection of the RNA pol I transcription; this detection does not necessarily need nucleolar disorganization. The data obtained with both permeabilized cells and microinjections of RNA precursors in the absence of glycerol support the hypothesis that the degree of hydration of the cell plays a role in RNA pol I transcription.
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PMID:Conditions favoring RNA polymerase I transcription in permeabilized cells. 866 Sep 46

A rice U3 small nuclear RNA (snRNA)-encoding gene has been isolated. The coding region of this gene contains all five conserved sequence boxes common to plant U3 snRNAs. The upstream and downstream regions of the gene harbour characteristic sequence elements required for transcription with RNA polymerase III (pol III), as well as three monocot-specific promoter (MSP) elements [Connelly et al., Mol. Cell Biol.14 (1994) 5910-5919], two of them comprising a palindromic G+C-rich segment. The sequence (TTTAAAA) of the TATA-box in this gene does not fit the established consensus [Marshallsay et al., Plant Mol. Biol. 19(1992) 973-983], making this gene unique among reported snRNA-encoding genes of plants. An RNase protection assay showed that the gene is expressed properly in cucumber protoplasts. We thus suggest that, with other promoter elements present, the TATA-box for RNA pol III-specific snRNA-encoding genes may not be as well conserved as that for RNA pol II-specific genes.
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PMID:Cloning, characterization and transient expression of the gene encoding a rice U3 small nuclear RNA. 868 6

A novel RNase activity was identified in a yeast RNA polymerase I (pol I) in vitro transcription system. Transcript cleavage occurred at the 3' end and was dependent on the presence of ternary pol I/DNA/RNA complexes and an additional protein factor not identical to transcription factor IIS (TFIIS). Transcript cleavage was observed both on arrested complexes at the linearized ends of the transcribed DNA and on intrinsic blocks of the DNA template. Shortened transcripts that remained associated within the ternary complexes were capable of resuming RNA chain elongation. Possible functions of the nuclease for transcript elongation or termination are discussed.
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PMID:A novel RNA polymerase I-dependent RNase activity that shortens nascent transcripts from the 3' end. 891 19

The 12 bp stem of the RNA hairpin responsible for the gag-pol frameshifting of the ribosomes during translation of the polycistronic HIV-1 mRNA has a pyrimidine-rich 5' strand and, consequently, a purine-rich 3' strand. Electrophoretic mobility shift assays have shown that DNA oligopyrimidines, 12 and 20 nucleotides long (but not oligopurines or G,T-containing oligomers), designed to form triplexes actually bind to the double-stranded RNA target. RNase V1 footprinting studies have confirmed the interaction between the hairpin stem and the RNA and 2'-O-methyl oligoribonucleotide analogues of the 12-mer oligodeoxypyrimidine as well as 5 propynyl,cytosine, containing the 12-mer oligodeoxypyrimidine, bind more strongly to the RNA target than the unmodified parent DNA oligomer. The complexes formed by the RNA hairpin and either the 12-mer oligodeoxypyrimidine or the 20-mer oligopyrimidine are stable at a neutral pH and in the absence of Mg2+ but blocked neither the reverse transcription nor cell-free translation of a RNA template in which the gag-pol frameshifting hairpin was inserted at the 5' end of the luciferase open reading frame.
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PMID:Binding of oligopyrimidines to the RNA hairpin responsible for the ribosome gag-pol frameshift in HIV-1. 1033 25

A model system to study the pathogenesis of gammaherpesvirus infections is the infection of mice with murine gammaherpesvirus 68 (MHV-68). To define the kinetics of infection, we developed an RNase protection assay to quantitate gene expression from lytic (K3, Rta, M8, DNA polymerase [DNA pol], and gB) and candidate latency (M2, M3, M9, M11, ORF73, and ORF74) genes. All candidate latency genes were expressed during lytic infection of 3T3 cells. Four kinetic classes of transcripts were observed following infection of 3T3 cells: immediate-early (K3, Rta, M8, and ORF73), early (DNA pol), early-late (M3, M11, and ORF74), and late (M2, M9, and gB). To assess the kinetics of viral gene expression in vivo, lungs, spleens, and mediastinal lymph nodes (MLN) were harvested from MHV-68-infected mice. All transcripts were expressed between 3 and 6 days postinfection (dpi) in the lungs. In the spleen, K3, M3, M8, and M9 transcripts were expressed between 10 and 16 dpi when latency is established. The K3, M3, M8, M9, and M11 transcripts were detected in the MLN from 2 through 16 dpi. This is the first demonstration of MHV-68 gene expression in the MLN. Importantly, our data showed that MHV-68 has different kinetics of gene expression at different sites of infection. Furthermore, we demonstrated that K3, a gene recently shown to encode a protein that downregulates major histocompatibility complex class I on the surface of cells, is expressed during latency, which argues for a role of K3 in immune evasion during latent infection.
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PMID:Kinetics of murine gammaherpesvirus 68 gene expression following infection of murine cells in culture and in mice. 1133 74

Feline immunodeficiency virus (FIV) is a lentivirus that causes AIDS-like immunodeficiency disease in domestic cats. Free-ranging lions, Panthera leo, carry a chronic species-specific strain of FIV, FIV-Ple, which so far has not been convincingly connected with immune pathology or mortality. FIV-Ple, harboring the three distinct strains A, B, and C defined by pol gene sequence divergences, is endemic in the large outbred population of lions in the Serengeti ecosystem in Tanzania. Here we describe the pattern of variation in the three FIV genes gag, pol-RT, and pol-RNase among lions within 13 prides to assess the occurrence of FIV infection and coinfection. Genome diversity within and among FIV-Ple strains is shown to be large, with strain divergence for each gene approaching genetic distances observed for FIV between different species of cats. Multiple in fections with two or three strains were found in 43% of the FIV-positive individuals based on pol-RT sequence analysis, which may suggest that antiviral immunity or interference evoked by one strain is not consistently protective against infection by a second. This comprehensive study of FIV-Ple in a free-ranging population of lions reveals a dynamic transmission of virus in a social species that has historically adapted to render the virus benign.
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PMID:Patterns of feline immunodeficiency virus multiple infection and genome divergence in a free-ranging population of African lions. 1501 97


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