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)

Overlapping cDNAs representing the complete L segment of Rift Valley fever virus were assembled, and the L protein was expressed via a recombinant vaccinia virus. The transcriptase activity of the L protein was assayed with two types of templates: natural ribonucleoproteins (RNPs) and artificial genome-like RNAs. RNPs purified in a CsCl gradient did not retain the RNA polymerase function, but the activity was restored when the L cDNA was expressed in mammalian cells via a recombinant vaccinia virus. Indeed, after transfection of transcriptase-depleted RNPs in cells infected with the recombinant vaccinia virus expressing the L protein, the mRNAs coding for the N and NSs proteins and to a lesser extent, those coding for the glycoproteins were synthesized as well as the corresponding proteins. The transcriptase activity of the recombinant L protein was then investigated by using synthetic templates containing the reporter chloramphenicol acetyltransferase gene in the antisense orientation flanked by the 3' and 5' noncoding region of the S genomic segment. Our results indicate that after transfection of the RNA templates, transcription was achieved in cells coexpressing both the L and N proteins. Together, the experiments demonstrate that the two proteins N and L are absolutely required and sufficient to reconstitute the transcriptase activity.
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PMID:The L protein of Rift Valley fever virus can rescue viral ribonucleoproteins and transcribe synthetic genome-like RNA molecules. 776 55

The complete nucleotide sequence of Toscana (TOS) virus (Bunyaviridae, Phlebovirus) L segment was determined. The L segment is 6404 nucleotides long, containing a single open reading frame (ORF) in the viral complementary sense coding for a protein of 2095 amino acids that, as in the case of negative strand RNA viruses, could be part of the RNA polymerase of TOS virus. This ORF is expressed by a messenger RNA (mRNA) as long as the genomic segment. Like the mRNAs expressed by the genomic segments of the other Bunyaviruses, the L mRNA has non-templated sequences at the 5' end. The comparison of TOS L protein sequence with the corresponding sequences of other negative strand RNA viruses showed a very high homology only with the Rift Valley Fever (RVF) virus. The residues conserved between the two proteins are mainly concentrated in the central region and contain three DD motifs proposed by Argos (1988) to be functional domains of DNA and RNA polymerases. The complete sequence of the Toscana virus L genomic segment has been deposited in the EMBL library with the accession number X68414.
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PMID:Toscana virus genomic L segment: molecular cloning, coding strategy and amino acid sequence in comparison with other negative strand RNA viruses. 846 May 26

A transcription system for Toscana virus (TOSV) (a member of the family BUNYAVIRIDAE:, genus PHLEBOVIRUS:) was constructed. For in vivo expression, the TOSV transcription system uses the viral N and L proteins and an S-like RNA genome containing the chloramphenicol acetyltransferase reporter gene in the antisense orientation flanked by the viral genomic 5'- and 3'-terminal S sequences. It was found that the N and L proteins represent the minimal protein requirement for an active transcription complex. To investigate the possibility of reassortment between TOSV and Rift Valley fever virus (RVFV), the activity of their polymerase complexes was tested on their heterologous S-like RNA genomes and this showed that both virus complexes were active. Moreover, hybrid transcriptase complexes with protein components originating from the two viruses were tested on both virus templates and only the combination RVFV L + TOSV N on RVFV S-like RNA was found to be active in this assay. These results suggest that virus reassortants might be generated whenever the two viruses infect the same host.
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PMID:Activity of Toscana and Rift Valley fever virus transcription complexes on heterologous templates. 1125 82

Reverse transcriptase PCR (RT-PCR) for diagnosis of Rift Valley fever (RVF) was evaluated by using 293 human and animal sera sampled during an RVF outbreak in Mauritania in 1998. Results of the RT-PCR diagnostic method were compared with those of virus isolation (VI) and detection of immunoglobulin M (IgM) antibodies. Our results showed that RT-PCR is a specific, sensitive tool for RVF diagnosis in the early phase of the disease and that its results do not differ significantly from those obtained by VI. Moreover, the combined results of RT-PCR and IgM antibody detection were in 100% concordance with the results of VI.
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PMID:Use of reverse transcriptase PCR in early diagnosis of Rift Valley fever. 1198 83

In the February 20 issue of Cell, report that Rift Valley Fever Virus (RVFV) targets cellular transcriptional apparatus to inhibit RNA polymerase II-mediated transcription. Unlike polio and vesicular stomatitis viruses, both of which target the TATA binding protein (TBP), RVFV appears to target the basal transcription factor THIIH to induce shut-off of host cell transcription.
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PMID:Targeting TFIIH to inhibit host cell transcription by Rift Valley Fever Virus. 1499 16

Rift Valley fever virus (RVFV) causes massive mosquito-borne epidemics among humans and decimates ruminants in which the mortality rate is about 1% and 10-30%, respectively. Morbidity in RVFV-infected humans is high largely due to the effects of hemorrhagic fever and encephalitis. This virus is native to sub-Saharan Africa; yet if this virus is introduced into the environment, virus transmission appears to occur whenever sheep and cattle are present with abundant mosquito populations. RVFV is a negative-strand RNA virus which belongs to the family Bunyaviridae, genus Phlebovirus, and contains tripartite-segmented genomes (S, M, and L). S-segment is the ambisense genome, where N and NSs genes are coded in an antiviral-sense and viral sense S-segment, respectively. The inhibition of host mRNA synthesis, which is induced by the binding of NSs protein to RNA polymerase II transcription factor TFIIH, is the primary reason for the host-protein shut-off in RVFV-infected cells. Development of a RVFV reverse genetics system, which has not been accomplished yet, is important for the study of viral replication mechanisms, host virus interaction, viral pathogenicity as well as vaccine evaluation and development.
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PMID:[Rift Valley fever virus]. 1574 61

Rift Valley fever virus (RVFV), which belongs to the genus Phlebovirus, family Bunyaviridae, has a tripartite negative-strand genome (S, M, and L segments) and is an important mosquito-borne pathogen for domestic animals and humans. We established an RVFV T7 RNA polymerase-driven minigenome system in which T7 RNA polymerase from an expression plasmid drove expression of RNA transcripts for viral proteins and minigenome RNA transcripts carrying a reporter gene between both termini of the M RNA segment in 293T cells. Like other viruses of the Bunyaviridae family, replication and transcription of the RVFV minigenome required expression of viral N and L proteins. Unexpectedly, the coexpression of an RVFV nonstructural protein, NSs, with N and L proteins resulted in a significant enhancement of minigenome RNA replication. Coexpression of NSs protein with N and L proteins also enhanced minigenome mRNA transcription in the cells expressing viral-sense minigenome RNA transcripts. NSs protein expression increased the RNA replication of minigenomes that originated from S and L RNA segments. Enhancement of minigenome RNA synthesis by NSs protein occurred in cells lacking alpha/beta interferon (IFN-alpha/beta) genes, indicating that the effect of NSs protein on minigenome RNA replication was unrelated to a putative NSs protein-induced inhibition of IFN-alpha/beta production. Our finding that RVFV NSs protein augmented minigenome RNA synthesis was in sharp contrast to reports that Bunyamwera virus (genus Bunyavirus) NSs protein inhibits viral minigenome RNA synthesis, suggesting that RVFV NSs protein and Bunyamwera virus NSs protein have distinctly different biological roles in viral RNA synthesis.
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PMID:Rift valley fever virus nonstructural protein NSs promotes viral RNA replication and transcription in a minigenome system. 1582 75

Rift Valley fever virus (RVFV) (genus Phlebovirus, family Bunyaviridae) has a tripartite negative-strand genome, causes a mosquito-borne disease that is endemic in sub-Saharan African countries and that also causes large epidemics among humans and livestock. Furthermore, it is a bioterrorist threat and poses a risk for introduction to other areas. In spite of its danger, neither veterinary nor human vaccines are available. We established a T7 RNA polymerase-driven reverse genetics system to rescue infectious clones of RVFV MP-12 strain entirely from cDNA, the first for any phlebovirus. Expression of viral structural proteins from the protein expression plasmids was not required for virus rescue, whereas NSs protein expression abolished virus rescue. Mutants of MP-12 partially or completely lacking the NSs open reading frame were viable. These NSs deletion mutants replicated efficiently in Vero and 293 cells, but not in MRC-5 cells. In the latter cell line, accumulation of beta interferon mRNA occurred after infection by these NSs deletion mutants, but not after infection by MP-12. The NSs deletion mutants formed larger plaques than MP-12 did in Vero E6 cells and failed to shut off host protein synthesis in Vero cells. An MP-12 mutant carrying a luciferase gene in place of the NSs gene replicated as efficiently as MP-12 did, produced enzymatically active luciferase during replication, and stably retained the luciferase gene after 10 virus passages, representing the first demonstration of foreign gene expression in any bunyavirus. This reverse genetics system can be used to study the molecular virology of RVFV, assess current vaccine candidates, produce new vaccines, and incorporate marker genes into animal vaccines.
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PMID:Rescue of infectious rift valley fever virus entirely from cDNA, analysis of virus lacking the NSs gene, and expression of a foreign gene. 1650 Nov 2

Rift Valley fever (RVF) virus belongs to the Bunyaviridae family of segmented negative-strand RNA viruses and causes mosquito-borne disease in sub-Saharan Africa. We report the development of a T7 RNA polymerase-driven plasmid-based genetic system for the virulent Egyptian isolate, ZH501. We have used this system to rescue a virus that has a 387 nucleotide deletion on the genomic M segment that eliminates the coding region for two non-structural proteins known as NSm. This virus, DeltaNSm rZH501, is indistinguishable from the parental ZH501 strain with respect to expression of structural proteins and growth in cultured mammalian cells.
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PMID:The NSm proteins of Rift Valley fever virus are dispensable for maturation, replication and infection. 1707 Aug 83

Rift Valley fever virus (RVFV, Bunyaviridae, Phlebovirus) is a mosquito-transmitted arbovirus that causes human and animal diseases in sub-Saharan Africa and was introduced into the Arabian Peninsula in 2000. Here, we describe a method of reverse genetics to recover infectious RVFV from transfected plasmids based on the use of the cellular RNA polymerase I promoter to synthesize viral transcripts. We compared its efficiency with a system using T7 RNA polymerase and found that both are equally efficient for the rescue of RVFV generating titers of approx. 10(7) to 10(8) pfu/ml. We used the RNA polymerase I-based system to rescue both attenuated MP12 and virulent ZH548 strains as well as chimeric MP12-ZH548 viruses, and in addition RVFV expressing reporter proteins.
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PMID:RNA polymerase I-mediated expression of viral RNA for the rescue of infectious virulent and avirulent Rift Valley fever viruses. 1861 93


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