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)

Field male Aedes aegypti (L.) and Aedes albopictus (Skuse) adults caught from fixed monitoring stations weekly for 1 yr were screened for dengue viruses (DEN-1, DEN-2, DEN-3, and DEN-4). The assay was carried out using a single-step reverse transcription (or transcriptase)-polymerase chain reaction (PCR) (RT-PCR) followed by a semi-nested PCR using an upstream consensus primer and four type-specific primers within the nonstructural protein three gene (NS3) of dengue viruses. The diagnostic fragments for DEN-1, DEN-2, DEN-3, and DEN-4 serotypes were of sizes 169, 362, 265, and 426 bp, respectively. Results showed that in Singapore 1.33% and 2.15% of Aedes aegypti and Aedes albopictus adult male mosquitoes, respectively, were positive for dengue viruses. The serotypes detected in male Ae. aegypti was DEN-1 (44%), followed by DEN-2 (22.2%) and DEN-3 (22.2%), and DEN-4 (11.1%). For Aedes albopictus males, the serotype was DEN-4 (38.9%), followed by DEN-2 (33.3%), DEN-3 (16.7%), and DEN-1 (11.1%).
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PMID:Detection of dengue viruses in field caught male Aedes aegypti and Aedes albopictus (Diptera: Culicidae) in Singapore by type-specific PCR. 1147 26

Hepatitis C virus (HCV) infection is usually treated with the combination of interferon and ribavirin, but only a small fraction of patients develop a sustained remission. There is need for the development of specific molecular approaches for the treatment of chronic HCV infection. We propose that RNA interference is highly effective antiviral strategy that offers great potential for the treatment of HCV infection. Three plasmid constructs expressing small interfering RNAs (siRNAs) targeted to sequences encoding the structural gene (E2) and non-structural genes (NS3, NS5B) of HCV1a genome were prepared. Antiviral properties of siRNAs against the HCV1a strain were studied in a transient replication model that involved the use of a transcription plasmid containing the full-length HCV genome and an adenovirus expressing T7 RNA polymerase. We found that siRNAs targeted to the E2, NS3 and NS5B regions of the HCV genome efficiently inhibited expression of the HCV core and NS5A protein measured by Western blot analysis and immunocytochemical staining. Intracytoplasmic immunization of siRNAs in HCV-transfected cells efficiently degraded genomic positive strand HCV RNA, as shown by ribonuclease protection assay (RPA). All three siRNAs efficiently inhibited synthesis of replicative negative strand HCV RNA in the transfected cells. A control siRNA plasmid against a Epstein--Barr virus latency gene did not inhibit protein expression and negative strand HCV RNA. These results suggest that RNAi is an effective and alternative approach that can be used to inhibit HCV expression and replication.
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PMID:Small interfering RNA effectively inhibits protein expression and negative strand RNA synthesis from a full-length hepatitis C virus clone. 1597 38

Since its identification in 1989, hepatitis C virus has been the subject of extensive research. The biology of the virus and the development of antiviral drugs are closely related. The RNA polymerase activity of nonstructural protein 5B was first demonstrated in 1996. NS5B is believed to localize to the perinuclear region, forming a replicase complex with other viral proteins. It has a typical polymerase structure with thumb, palm, and finger domains encircling the active site. A de novo replication initiation mechanism has been suggested. To date, many small molecule inhibitors are known including nucleoside analogues, non-nucleoside analogues, and pyrophosphate mimics. NS5B interacts with other viral proteins such as core, NS3, 4A, 4B, and 5A. The helicase activity of NS3 seems necessary for RNA strand unwinding during replication, with other nonstructural proteins performing modulatory roles. Cellular proteins interacting with NS5B include VAMP-associated proteins, heIF4AII, hPLIC1, nucleolin, PRK2, a-actinin, and p68 helicase. The interactions of NS5B with these proteins might play roles in cellular trafficking, signal transduction, and RNA polymerization, as well as the regulation of replication/translation processes.
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PMID:Nonstructural protein 5B of hepatitis C virus. 1681 94

The nonstructural protein NS2-3 of pestiviruses undergoes tightly regulated processing. For bovine viral diarrhea virus it was shown that uncleaved NS2-3 is required for infectious particle formation while cleaved NS3 is essential for genome replication. To further investigate the functions of NS2-3 and NS4A in the pestivirus life cycle, we established T7 RNA polymerase-dependent trans-complementation for p7-NS2-3-4A of classical swine fever virus (CSFV). Expression of NS2-3 and NS4A in trans restored the production of infectious particles from genomes lacking NS2-3 expression. Co-expression of cleaved NS4A was essential. None of the enzymatic activities harbored by NS2-3 were required for infectious particle formation. Importantly, expression of uncleavable NS2-3 together with NS4A rescued infectious particles from a genome lacking NS2, demonstrating that cleaved NS2 per se has no additional essential function. These data indicate that NS2-3 and NS3, each in association with NS4A, have independent functions in the CSFV life cycle.
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PMID:Nonstructural proteins NS2-3 and NS4A of classical swine fever virus: essential features for infectious particle formation. 1748 32

The hepatitis C Virus (HCV) presents a high degree of genetic variability which is explained by the combination of a lack of proof reading by the RNA dependant RNA polymerase and a high level of viral replication. The resulting genetic polymorphism defines a classification in clades, genotypes, subtypes, isolates and quasispecies. This diversity is known to reflect the range of responses to Interferon therapy. The genotype is one of the predictive parameters currently used to define the antiviral treatment strategy and the chance of therapeutic success. Studies have also reported the potential impact of the viral genetic polymorphism in the outcome of antiviral therapy in patients infected by the same HCV genotype. Both structural and non structural genomic regions of HCV have been suggested to be involved in the Interferon pathway and the resistance to antiviral therapy. In this review, we first detail the viral basis of HCV diversity. Then, the HCV genetic regions that may be implicated in resistance to therapy are described, with a focus on the structural region encoded by the E2 gene and the non-structural genes NS3, NS5A and NS5B. Both mechanisms of the Interferon resistance and of the new antiviral drugs are described in this review.
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PMID:Genetic diversity of the hepatitis C virus: impact and issues in the antiviral therapy. 1755 24

The next generation of anti-HCV therapeutics agents will fall into new interferons and interferon inducers, alternatives to ribavirin, specific HCV inhibitors and immune therapies. According to HCV variability, the clinical success of HCV-targeted drugs will depend on their ability to suppress all viral variants as well as prevent the emergence of resistant viruses. Examination of these properties are limited in the absence of convenient animal model of HCV infection and limited possibilities studies on HCV replicons. The NS3-4A serine protease and the NS5B RNA polymerase have emerged as popular target for antiviral intervention. Blockage ofNS3 activity is expected to inhibit HCV replication by direct suppression of viral protein production and restoration of host responsiveness to IFN. The experience with synthetic agonists of Toll-like receptors 7 and 9 have begun to show their potential in controlling HCV infection. The new anti-HCV agents have progressed through early-phase clinical trials. Based on HCV infection history it seems that IFN will be the gold standard of the therapy and new agents probably will administer in combination with IFN.
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PMID:[Treatment of chronic hepatitis C--new drugs, new hope]. 1768 54

Hepatitis C virus (HCV) infects over 170 million persons worldwide. It is the leading cause of liver disease in the U.S. and is responsible for most liver transplants. Current treatments for this infectious disease are inadequate; therefore, new therapies must be developed. Several labs have obtained evidence for a protein complex that involves many of the nonstructural (NS) proteins encoded by the virus. NS3, NS4A, NS4B, NS5A, and NS5B appear to interact structurally and functionally. In this study, we investigated the interaction between the helicase, NS3, and the RNA polymerase, NS5B. Pull-down experiments and surface plasmon resonance data indicate a direct interaction between NS3 and NS5B that is primarily mediated through the protease domain of NS3. This interaction reduces the basal ATPase activity of NS3. However, NS5B stimulates product formation in RNA unwinding experiments under conditions of excess nucleic acid substrate. When the concentrations of NS3 and NS5B are in excess of nucleic acid substrate, NS5B reduces the rate of NS3-catalyzed unwinding. Under pre-steady-state conditions, in which NS3 and substrate concentrations are similar, product formation increased in the presence of NS5B. The increase was consistent with 1:1 complex formed between the two proteins. A fluorescently labeled form of NS3 was used to investigate this interaction through fluorescence polarization binding assays. Results from this assay support interactions that include a 1:1 complex formed between NS3 and NS5B. The modulation of NS3 by NS5B suggests that these proteins may function together during replication of the HCV genome.
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PMID:RNA unwinding activity of the hepatitis C virus NS3 helicase is modulated by the NS5B polymerase. 1817 52

A new class of compounds--acridone derivatives--was tested using the direct fluorometric helicase activity assay to determine the inhibitory properties of the derivatives towards the NS3 helicase of Hepatitis C virus (HCV). The compounds were also tested as putative transcription inhibitors of in vitro transcription based on the DNA-dependent T7 RNA polymerase. Most of the acridone derivatives tested were transcription inhibitors; however, only four of them inhibited the NS3 helicase at low concentrations (IC(50) from 3 microM to 20 microM) and were therefore selected for further studies on the mechanism of inhibition. The acridone derivatives probably act via intercalation into double-stranded nucleic acids but they may also interact directly with viral enzymes. Selected carboxamides were tested in the subgenomic HCV replicon system. Two of the compounds: N-(pyridin-4-yl)-amide and N-(pyridin-2-yl)-amide of acridone-4-carboxylic acid are efficient RNA replication inhibitors with selectivity indexes of 19.4 and 40.5, respectively, proving that the acridone derivatives may be regarded as potential antiviral agents.
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PMID:New acridone-4-carboxylic acid derivatives as potential inhibitors of hepatitis C virus infection. 1880 60

The non-structural protein NS1 of Periplaneta fuliginosa densovirus (PfDNV) is a multifunctional protein that has previously been shown to possess ATP-binding, ATPase, site-specific DNA-binding, helicase, and transcription activation activities. We report here an investigation of the cytopathogenicity of this viral non-structural (NS) protein, as well as other two NSs, NS2, and NS3, in cultured insect cells. The expression of NS1 alone potently inhibited cellular gene expression, whereas NS2 and NS3 did not produce a similar effect. The inhibition of gene expression by NS1 was confirmed to be specific and not a simple manifestation of toxicity. For example, NS1 inhibited expression of several reporter genes under the control of different RNA polymerase II promoters, whereas it did not inhibit expression from a T7 RNA polymerase promoter construct. Mapping analysis identified the carboxy-terminal peptide of this protein as the region important for the inhibition of cellular gene expression, suggesting that this inhibition is independent of its DNA-binding activity. Next, the mutagenesis assay showed that ATP-binding was essential for the unique function of this protein. Furthermore, we found that NS2 and NS3 cooperatively enhanced the NS1-induced transcription inhibition. Co-expression of all the three NS proteins in Sf9 cells also led to necrotic cell death by ATP depletion.
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PMID:Non-structural proteins of Periplaneta fuliginosa densovirus inhibit cellular gene expression and induce necrosis in Sf9 cell cultures. 1929 99

Inter-species transmission is often incriminated in the epidemiology of Pestivirus diseases. The purpose of this study was to investigate the prevalence of Pestivirus in some mountain wild ungulates and to determine their role in Pestivirus transmission, as mountain pastures are a place where cohabitations between wild and domestic ungulates are particularly high. Between 2003 and 2007, a longitudinal epidemiological study was carried out on hunted ungulates in the French Hautes-Alpes department. Pestivirus-specific antibodies against p80 protein (also named NS3) common to all Bovine Viral Diarrhea Virus (BVDV) and Border Disease Virus (BDV) were found in 45.9% (95% confidence interval [CI95%]: 40.5-51.3%) of the 343 tested chamois (Rupicapra rupicapra). In addition, mouflons (Ovis gmelinii musimon) were shown for the first time to be strongly infected (61.1%; CI95%: 38.6-83.6) by a Pestivirus. These serological ELISA results were confirmed by comparative virus neutralization tests, performed on seven Pestivirus strains by using 15 seropositive samples. The highest antibody titers were directed against 2 BDV strains (Av and 33s strains), rather than BDV-4, a strain responsible for Pyrenean-chamois epizooties. Virus neutralization tests confirm a BDV circulation in wild ungulates in the French South Alps. However, no Pestivirus RNA was detected by reverse-transcriptase polymerase chain reaction in serum and spleen samples from seronegative animals and no virus was isolated from those samples either. Efforts should be made to improve the protocol in order to be able to isolate and characterize the local strain. Finally, the oldness (age) and femaleness (gender) increase the risk of seroconversion in chamois.
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PMID:Epidemiology of Pestivirus infection in wild ungulates of the French South Alps. 2070 72


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