In vivo reconstitution of active Thogoto virus polymerase: assays for the compatibility with other orthomyxovirus core proteins and template RNAs.
Tick-borne Thogoto virus (THOV), the prototype of a new genus in the Orthomyxoviridae family, contains six single-stranded RNA segments of negative polarity. Four of them encode gene products that correspond to the influenza virus PB1, PB2, PA and NP core proteins. Here we describe an in vivo system in which the expression of a THOV model RNA is driven by THOV core proteins synthesized from cloned cDNAs. Our results demonstrated the biological activity of our cloned genes and showed that the three polymerase subunits and the NP are required for gene expression. For comparison, we also used the in vivo reconstituted systems of the influenza A and B viruses. None of the polymerase or NP proteins was active in a heterologous orthomyxovirus core, indicating a high specificity in core assembly and/or function. Interestingly, the THOV polymerase did not recognize the influenza A virus promoter and vice versa.
Weber
F
F
Department of Virology, University of Freiburg, Germany.
Jambrina
E
E
González
S
S
Dessens
J T
JT
Leahy
M
M
Kochs
G
G
Portela
A
A
Nuttall
P A
PA
Haller
O
O
Ortín
J
J
Zürcher
T
T
D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000378:metabolism*; D015967:Gene Expression Regulation, Viral; D009975:Orthomyxoviridae / Q000235:genetics*; D012367:RNA, Viral / Q000235:genetics; D016865:Reassortant Viruses; D013698:Templates, Genetic; D018117:Thogotovirus / Q000201:enzymology* / Q000235:genetics; D014758:Viral Core Proteins / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*
D012367:RNA, Viral; D014758:Viral Core Proteins; D014764:Viral Proteins; D012321:DNA-Directed RNA Polymerases