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dc.contributor.author
Iglesias, Nestor Gabriel
dc.contributor.author
Filomatori, Claudia Veronica
dc.contributor.author
Gamarnik, Andrea Vanesa
dc.date.available
2017-02-08T21:56:48Z
dc.date.issued
2011-06
dc.identifier.citation
Iglesias, Nestor Gabriel; Filomatori, Claudia Veronica; Gamarnik, Andrea Vanesa; F1 Motif of Dengue Virus Polymerase NS5 Is Involved in Promoter-Dependent RNA Synthesis; American Society For Microbiology; Journal Of Virology; 85; 12; 6-2011; 5745-5756
dc.identifier.issn
0022-538X
dc.identifier.uri
http://hdl.handle.net/11336/12784
dc.description.abstract
The mechanism by which viral RNA-dependent RNA polymerases (RdRp) specifically amplify viral genomes is still unclear. In the case of flaviviruses, a model has been proposed that involves the recognition of an RNA element present at the viral 5' untranslated region, stem-loop A (SLA), that serves as a promoter for NS5 polymerase binding and activity. Here, we investigated requirements for specific promoter-dependent RNA synthesis of the dengue virus NS5 protein. Using mutated purified NS5 recombinant proteins and infectious viral RNAs, we analyzed the requirement of specific amino acids of the RdRp domain on polymerase activity and viral replication. A battery of 19 mutants was designed and analyzed. By measuring polymerase activity using nonspecific poly(rC) templates or specific viral RNA molecules, we identified four mutants with impaired polymerase activity. Viral full-length RNAs carrying these mutations were found to be unable to replicate in cell culture. Interestingly, one recombinant NS5 protein carrying the mutations K456A and K457A located in the F1 motif lacked RNA synthesis dependent on the SLA promoter but displayed high activity using a poly(rC) template. Promoter RNA binding of this NS5 mutant was unaffected while de novo RNA synthesis was abolished. Furthermore, the mutant maintained RNA elongation activity, indicating a role of the F1 region in promoter-dependent initiation. In addition, four NS5 mutants were selected to have polymerase activity in the recombinant protein but delayed or impaired virus replication when introduced into an infectious clone, suggesting a role of these amino acids in other functions of NS5. This work provides new molecular insights on the specific RNA synthesis activity of the dengue virus NS5 polymerase.
dc.format
application/pdf
dc.language.iso
eng
dc.publisher
American Society For Microbiology
dc.rights
info:eu-repo/semantics/restrictedAccess
dc.rights.uri
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.subject
Dengue Virus
dc.subject
Rna Synthesis
dc.subject
Viral Rna Polymerases
dc.subject.classification
Virología
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Ciencias Biológicas
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CIENCIAS NATURALES Y EXACTAS
dc.title
F1 Motif of Dengue Virus Polymerase NS5 Is Involved in Promoter-Dependent RNA Synthesis
dc.type
info:eu-repo/semantics/article
dc.type
info:ar-repo/semantics/artículo
dc.type
info:eu-repo/semantics/publishedVersion
dc.date.updated
2017-02-07T13:52:25Z
dc.journal.volume
85
dc.journal.number
12
dc.journal.pagination
5745-5756
dc.journal.pais
Estados Unidos
dc.journal.ciudad
Washington
dc.description.fil
Fil: Iglesias, Nestor Gabriel. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina
dc.description.fil
Fil: Filomatori, Claudia Veronica. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina
dc.description.fil
Fil: Gamarnik, Andrea Vanesa. Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Bioquimicas de Buenos Aires; Argentina. Fundación Instituto Leloir; Argentina
dc.journal.title
Journal Of Virology
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/url/http://jvi.asm.org/content/85/12/5745.long
dc.relation.alternativeid
info:eu-repo/semantics/altIdentifier/doi/http://dx.doi.org/10.1128/JVI.02343-10


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