Specific requirements for elements of the 5' and 3' terminal regions in flavivirus RNA synthesis and viral replication.
We initially studied requirements for 5' and 3' terminal regions (TRs) in flavivirus negative strand synthesis in vitro. Purified West Nile (WNV) and dengue-2 (DV2) RNA polymerases were both active with all-WNV or all-DV2 subgenomic RNAs containing the 5'- and 3'TRs of the respective genomes. However, subgenomic RNAs in which the 5'-noncoding region (5'NCR) or the 5'ORF (nts 100-230) in the 5'TR were substituted by analogous sequences derived from the heterologous genome were modestly to severely defective as templates for either polymerase. We also evaluated the infectivity of substitution mutant WNV genome-length RNAs. All WNV RNAs containing the DV2 3'SL were unable to replicate. However, WNV RNAs containing substitutions of the 5'NCR, the capsid gene, and/or 3'NCR nt sequences upstream from the WNV 3'SL, by the analogous DV2 nt sequences, were infectious. Combined results suggested that replication was not dependent upon species homology between the 3'SL and NS5.
Yu
Li
L
Laboratory of Vector-Borne Virus Diseases, Division of Viral Products, Office of Vaccines Research and Review, CBER, FDA, Bethesda, Maryland, USA. yuli@cber.fda.gov
Nomaguchi
Masako
M
Padmanabhan
R
R
Markoff
Lewis
L
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D002522:Chlorocebus aethiops; D009033:Culicidae; D003716:Dengue Virus / Q000235:genetics* / Q000502:physiology; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D022661:RNA, Untranslated / Q000235:genetics*; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics*; D012324:RNA-Dependent RNA Polymerase / Q000378:metabolism; D012045:Regulatory Sequences, Nucleic Acid; D010948:Viral Plaque Assay; D014764:Viral Proteins / Q000378:metabolism; D014779:Virus Replication / Q000502:physiology*; D014902:West Nile virus / Q000235:genetics* / Q000502:physiology
D022661:RNA, Untranslated; D012367:RNA, Viral; D014764:Viral Proteins; D012324:RNA-Dependent RNA Polymerase