Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.1.27.5 (RNase)
 17,967 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Chemical instability of some of the phosphodiester bonds, often observed in large RNAs, visualizes the autocatalytic properties of this class of nucleic acids. Unexpectedly, selective hydrolysis occurs also in short oligoribonucleotides (as short as a tetramer or hexamer). Herein, we describe additional experiments which support the conclusion that the hydrolysis is not due to ribonuclease contamination but is of autocatalytic origin and is related to the sequence and structure of single-stranded oligomers. Moreover, we show that the presence in the reaction mixture of polyamines, such as spermidine, is essential for hydrolysis of oligoribonucleotides.
Acta Biochim Pol 1999
PMID:Nonenzymatic hydrolysis of oligoribonucleotides. V. The elements affecting the process of self-hydrolysis. 1045 90

The encapsidation signal of the yeast L-A virus contains a 24-nucleotide stem-loop structure with a 5-nucleotide loop and an A bulged at the 5' side of the stem. The Pol part of the Gag-Pol fusion protein is responsible for encapsidation of viral RNA. Opened empty viral particles containing Gag-Pol specifically bind to this encapsidation signal in vitro. We found that binding to empty particles protected the bulged A and the flanking-two nucleotides from cleavage by Fe(II)-EDTA-generated hydroxyl radicals. The five nucleotides of the loop sequence ((4190)GAUCC(4194)) were not protected. However, T1 RNase protection and in vitro mutagenesis experiments indicated that G(4190) is essential for binding. Although the sequence of the other four nucleotides of the loop is not essential, data from RNase protection and chemical modification experiments suggested that C(4194) was also directly involved in binding to empty particles rather than indirectly through its potential base pairing with G(4190). These results suggest that the Pol domain of Gag-Pol contacts the encapsidation signal at two sites: one, the bulged A, and the other, G and C bases at the opening of the loop. These two sites are conserved in the encapsidation signal of M1, a satellite RNA of the L-A virus.
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PMID:Recognition of RNA encapsidation signal by the yeast L-A double-stranded RNA virus. 1095 12

We have examined the influence of RNA upon the interaction of Gag-Pol with Gag during human immunodeficiency virus type 1 (HIV-1) assembly. COS7 cells were transfected with protease-negative HIV-1 proviral DNA, and Gag/Gag-Pol complexes were detected by coimmunoprecipitation with anti-integrase. In COS7 cells, Gag/Gag-Pol is found almost entirely in pelletable, membrane-bound complexes. Exposure of cells to 1% Triton X-100 releases Gag/Gag-Pol from bulk membrane, but the complexes remain pelletable. The role of RNA in facilitating the interaction between Gag and Gag-Pol was examined in these bulk membrane-free, pelletable complexes. The specific presence of viral genomic RNA is not required to maintain the Gag/Gag-Pol interaction, but some type of RNA is, since exposure to RNase destabilized the Gag/Gag-Pol complex. When present only in Gag, the nucleocapsid mutation R7R10K11S, which inhibits Gag binding to RNA, inhibits the formation of both Gag and Gag/Gag-Pol complexes. When present only in Gag-Pol, this mutation has no effect upon complex formation. This result indicates that Gag-Pol may not interact directly with RNA but rather requires RNA-facilitated Gag multimerization for its interaction with Gag.
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PMID:Role of RNA in facilitating Gag/Gag-Pol interaction. 1190 55

Potato leafroll virus is a member of the polerovirus genus. The isometric virion is formed by a coat protein encapsidating single-stranded, positive-sense, mono-partite genomic RNA with covalently attached viral protein at the 5' end. The coat protein of the virus exists in two forms: i) a 23 kDa protein, the product of the coat protein gene, and ii) a 78 kDa protein, the product of the coat protein gene and an additional open reading frame expressed by read-through of the coat protein gene stop codon. The aim of this work was the expression of potato leafroll virus coat protein-based proteins that would be able to assemble into virus-like particles in insect cells. These modified particles were tested for their ability to encapsidate nucleic acids. Two types of N-terminally His-tagged coat protein constructs were used for the expression in insect cells: one, encoding a 23 kDa protein with the C-terminal amino-acid sequence corresponding to the wild type coat protein and the second with additional clathrin binding domain at the C-terminus. The expression of these two proteins by a recombinant baculovirus was characterized by Western immunoblotting with antibodies directed against potato leafroll virus. The protection or putative encapsidation of nucleic acids by these two coat protein derivatives was shown by DNase I and RNase A protection assays.
Acta Biochim Pol 2005
PMID:Virus-like particles of potato leafroll virus as potential carrier system for nucleic acids. 1617 44

hnRNP K protein, which localizes to the nucleus, cytoplasm and mitochondria, is involved in the various cellular processes that compose gene expression. We used a SAGE-based assay to profile RNAs associated with hnRNP K protein in rat mitochondria. RNA was isolated from mitoplasts obtained from highly purified and RNase-treated mitochondria. Total RNA and RNA associated with hnRNP K protein were then used as input material for generating two SAGE libraries. Mitochondrion-derived tags isolated from the total mitoplast RNA library represented 86.3%, while those isolated from the library constructed from RNA associated with hnRNP K protein represented only 28.2% of selected tags. Thus, an unexpected number of nuclear-encoded RNAs were purified from mitochondria. Many of these transcripts were co-purified with hnRNP K protein, and high levels of nuclear-encoded RNAs co-immunoprecipitating with K protein corresponded to elevated hnRNP K protein levels of the organelle. The most abundant RNAs that were co-purified with hnRNP K protein represented transcripts originating from satellite I DNA. While satellite I RNA levels were higher in the nucleus and cytoplasm than in mitochondria, the most abundant binding of satellite I transcripts to hnRNP K protein was found in mitochondria. The role of satellite I RNA in mitochondria remains to be elucidated.
Acta Biochim Pol 2006
PMID:Mitochondria-associated satellite I RNA binds to hnRNP K protein. 1649 41

To characterize proteins associated with active transcription complexes, we purified RNA polymerase II (pol II) from Saccharomyces cerevisiae after fixing live cells with formaldehyde. The approach mimics ChIP and requires solubilizing cross-linked complexes with sonication. Pol II was affinity-purified, and associated proteins were identified by MS. Several classes of proteins depended on cross-linking, including Mediator, general transcription factors, elongation factors, ribonucleoprotein particle (RNP) proteins, and histones. A tagged RNP protein reciprocally purified pol II under identical cross-linking conditions, and the association between RNP proteins and pol II was largely RNase-sensitive. The data indicate that the cross-linked Pol II purification contains elongating pol II with associated nascent RNP. Consistent with this view, some elongation factors no longer associate with pol II after inactivation of transcription in the temperature-sensitive pol II mutant, rpb1-1. Taken together, our data suggest that the cross-linked pol II purification contains a mixed population of pol II, including initiating pol II and elongating pol II.
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PMID:Protein characterization of Saccharomyces cerevisiae RNA polymerase II after in vivo cross-linking. 1807 27

The Paf1 complex (Paf1, Ctr9, Cdc73, Rtf1, and Leo1) is normally associated with RNA polymerase II (Pol II) throughout the transcription cycle. However, the loss of either Rtf1 or Cdc73 results in the detachment of the Paf1 complex from Pol II and the chromatin form of actively transcribed genes. Using functionally tagged forms of the Paf1 complex factors, we have determined that, except for the more loosely associated Rtf1, the remaining components stay stably associated with one another in an RNase-resistant complex after dissociation from Pol II and chromatin. The loss of Paf1, Ctr9, or to a lesser extent Cdc73 or Rtf1 results in reduced levels of serine 2 phosphorylation of the Pol II C-terminal domain and in increased read through of the MAK21 polyadenylation site. We found that the cleavage and polyadenylation factor Cft1 requires the Pol II-associated form of the Paf1 complex for full levels of interaction with the serine 5-phosphorylated form of Pol II. When the Paf1 complex is dissociated from Pol II, a direct interaction between Cft1 and the Paf1 complex can be detected. These results are consistent with the Paf1 complex providing a point of contact for recruitment of 3'-end processing factors at an early point in the transcription cycle. The lack of this connection helps to explain the defects in 3'-end formation observed in the absence of Paf1.
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PMID:Direct interactions between the Paf1 complex and a cleavage and polyadenylation factor are revealed by dissociation of Paf1 from RNA polymerase II. 1846 35

RNA interference (RNAi)-based gene silencing possesses great therapeutic potential for inhibiting replication of human viruses such as hepatitis C virus (HCV). However, one of the putative limitations for its use as a therapy is the rapid emergence of escape variants. These contain deletions or mutations within the viral genome sequences complementary to the small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) being used for treatment. As a potential solution to this problem, we constructed an expression system for duplex RNAs harboring two siRNA units using convergent H1 and U6 Pol III promoters. Here, the length and orientation of the transcript, tandem siRNA (tsiRNA), were optimized to be processed by the intracellular ribonuclease Dicer into functional siRNAs targeting different sequences. Assessment in transfected cells indicates that the length of the tsiRNA duplex (40-42 base pairs) is more critical for both siRNA-producing capacity and gene silencing activity than the orientation of each siRNA unit. In Huh7 cells replicating full-length HCV RNA, expression of length-optimized tsiRNA inhibited viral protein levels as efficiently as a single 21-nucleotide siRNA-expression construct, without affecting miRNA maturation or induction of an interferon response. We verified that the anti-viral activity of tsiRNA was achieved by precise cleavage of two target sites. A distinct advantage of this strategy is that each side of the optimized linear duplex RNA could enter into the Dicer-mediated processing machinery, thus likely providing more equal and efficient production of multiple siRNAs required for reducing the chance of viral escape.
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PMID:Optimization of linear double-stranded RNA for the production of multiple siRNAs targeting hepatitis C virus. 1932 60

The SR proteins are a family of pre-mRNA splicing factors with additional roles in gene regulation. To investigate individual family members in vivo, we generated a comprehensive panel of stable cell lines expressing GFP-tagged SR proteins under endogenous promoter control. Recruitment of SR proteins to nascent FOS RNA was transcription dependent and RNase sensitive, with unique patterns of accumulation along the gene specified by the RNA recognition motifs (RRMs). In addition, all SR protein interactions with Pol II were RNA dependent, indicating that SR proteins are not preassembled with Pol II. SR protein interactions with RNA were confirmed in situ by FRET/FLIM. Interestingly, SC35-GFP also exhibited FRET with DNA and failed to associate with cytoplasmic mRNAs, whereas all other SR proteins underwent nucleocytoplasmic shuttling and associated with specific nuclear and cytoplasmic mRNAs. Because different constellations of SR proteins bound nascent, nuclear, and cytoplasmic mRNAs, mRNP remodeling must occur throughout an mRNA's lifetime.
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PMID:SR protein family members display diverse activities in the formation of nascent and mature mRNPs in vivo. 1939 95

The oligomerization of HIV-1 Gag and Gag-Pol proteins, which are assembled at the plasma membrane, leads to viral budding. The budding generally places the viral components under non-reducing conditions. Here the effects of non-reducing conditions on Gag structures and viral RNA protection were examined. Using different reducing conditions and SDS-PAGE, it was shown that oligomerized Gag possesses intermolecular covalent bonds under non-reducing conditions. In addition, it was demonstrated that the mature viral core contains a large amount of covalent bonded Gag multimers, as does the immature core. Viral genomic RNA becomes sensitive to ribonuclease in reducing conditions. These results suggest that, under non-reducing conditions, covalent bonded Gag multimers are formed within the viral particles and play a role in protection of the viral genome.
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PMID:Covalent bonded Gag multimers in human immunodeficiency virus type-1 particles. 1990 61


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