In vivo reconstitution of active Thogoto virus polymerase: assays for the compatibility with other orthomyxovirus core proteins and template RNAs

• Published in: Virus research Vol. 58; no. 1; pp. 13 - 20
• Main Authors: Weber, Friedemann, Jambrina, Enrique, González, Susana, Dessens, Johannes T, Leahy, Michael, Kochs, Georg, Portela, Agustı́n, Nuttall, Patricia A, Haller, Otto, Ortı́n, Juan, Zürcher, Thomas
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01-11-1998
• Subjects: CAT expression; Cloning, Molecular; DNA-Directed RNA Polymerases - metabolism; Gene Expression Regulation, Viral; In vivo transcription/replication system; Influenza A virus; Orthomyxoviridae - genetics; Orthomyxoviruses; Reassortant Viruses; RNA, Viral - genetics; Templates, Genetic; Thogoto virus; Thogotovirus - enzymology; Thogotovirus - genetics; Viral Core Proteins - genetics; Viral Proteins - genetics; Viral Proteins - metabolism; Orthomyxoviruses; Thogoto virus; In vivo transcription/replication system; CAT expression
• ISSN: 0168-1702; 1872-7492
• DOI: 10.1016/S0168-1702(98)00096-3

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.