title: [5´cap -independent translation of dengue virus genomic RNA].
abstract: OBJECTIVE
  To analyze the involvement of methyl guanosine triphosphate cap (5cap) and the start site of the genomic RNA of Dengue virus serotype 2 (DENV-2) American genotype in translation, using a cell-free system prepared from human placenta.
  
  
  METHODS
  The recombinant plasmid pTZ18R-D2 was prepared containing DNA encoding the 5UTR and the first 201 nucleotides of the viral capsid. This plasmid was used to transcribe the corresponding RNA (RNA-D2) without the 5 cap. The RNA-D2 was translated in a system consisting of the postmitochondrial fraction (S-30) from human placenta and the incorporation of [14C] aminoacids in the presence of RNA-D2 and in its absence (control) was evaluated. Seven antisense oligonucleotides (OAs1-7) directed against sequences of the SLA, SLB and CHP structures of RNA-D2 were designed and the effect thereof on RNA-D2 translation was analyzed.
  
  
  RESULTS
  The RNA-D2 produced a significant increase (p<0.001) in the incorporation of [14C] amino acids, with 75% stimulation of translational activity compared to the control. Analysis of the translation products showed peak incorporation corresponding to peptides with apparent molecular weight close to the expected (7.746 kDa).The OAs5, complementary to a sequence of SLB structure of RNA-D2, completely inhibited translation.
  
  
  CONCLUSIONS
  The RNA-D2 was translated specifically and efficiently under conditions similar to human intracellular conditions, by an alternative 5 cap-independent mechanism, which would involve the SLB structure. This mechanism might be seen as an aim in the development of antisense therapies to inhibit virus replication.
authors.affiliation: Facultad de Odontología, Universidad de Carabobo, Valencia, Venezuela.
authors.forename: Mariela
authors.initials: M
authors.lastname: Pérez Dominguez
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Roselbis
authors.initials: R
authors.lastname: Rojas
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Eduardo
authors.initials: E
authors.lastname: Bandeira
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Dayana
authors.initials: D
authors.lastname: Requena
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Ana C
authors.initials: AC
authors.lastname: Ferreras
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Juana L
authors.initials: JL
authors.lastname: Triana
authors.affiliation: Facultad de Ciencias de la Salud, Universidad de Carabobo, Maracay, Venezuela.
authors.forename: Francisco
authors.initials: F
authors.lastname: Triana-Alonso
mesh_terms: D003716:Dengue Virus / Q000235:genetics*; D023281:Genomics; D006801:Humans; D014176:Protein Biosynthesis; D012315:RNA Caps / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics*
chemical_list: D012315:RNA Caps; D012367:RNA, Viral