The three subunits of the polymerase and the nucleoprotein of influenza B virus are the minimum set of viral proteins required for expression of a model RNA template.
The genes encoding the nucleoprotein, PB1, PB2, and PA proteins of the influenza virus strain B/Panamá/45/90 have been cloned under control of the T7 RNA polymerase promoter of plasmid pGEM-3. Transfection of the recombinant plasmids obtained into mammalian cells, which had been infected with a vaccinia virus encoding the T7 RNA polymerase, resulted in expression of the expected influenza B virus polypeptides. Moreover, it is shown that coexpression of the four recombinant core proteins in COS-1 cells reconstituted a functional polymerase capable of expressing a synthetic influenza B virus-like CAT RNA. By using the influenza B virus recombinant plasmids and a set of pGEM-derived plasmids encoding the homologous core proteins of the influenza A virus A/Victoria/3/75 (I. Mena et al. (1994). J. Gen. Virol. 75, 2109-2114), the capabilities of homo- and heterotypic mixtures of the four core proteins to express synthetic type A and B CAT RNAs were analyzed. Both the influenza A and B virus polymerases were active in expressing, albeit with reduced efficiencies, the heterotypic model CAT RNAs. However, none of all possible heterotypic mixtures of the core proteins reconstituted a functional polymerase. In order to fully characterize the recombinant plasmids obtained, the nucleotide sequences of the cloned genes were determined and compared to sequences of other type B virus isolates. The results obtained from these latter analyses are discussed in terms of the conservation and evolution of the influenza B virus core genes.
Jambrina
E
E
Instituto de Salud Carlos III, Centro Nacional de Biología Fundamental, Majadahonda 28220, Madrid, Spain.
Bárcena
J
J
Uez
O
O
Portela
A
A
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D019556:COS Cells; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000096:biosynthesis / Q000235:genetics*; D009981:Influenza B virus / Q000235:genetics*; D008969:Molecular Sequence Data; D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; D012324:RNA-Dependent RNA Polymerase; D011994:Recombinant Proteins / Q000235:genetics; D014758:Viral Core Proteins / Q000096:biosynthesis / Q000235:genetics*; D014764:Viral Proteins / Q000096:biosynthesis / Q000235:genetics*
D019590:Nucleocapsid Proteins; D009698:Nucleoproteins; C061290:PA protein, influenza viruses; C057027:PB2 protein, Influenzavirus A; D011994:Recombinant Proteins; D014758:Viral Core Proteins; D014764:Viral Proteins; C077346:influenza virus polymerase basic protein 1; D012324:RNA-Dependent RNA Polymerase; D012321:DNA-Directed RNA Polymerases