Flexibility of bunyavirus genomes: creation of an orthobunyavirus with an ambisense S segment

Journal of Virology
Ingeborg van Knippenberg, Richard M Elliott

Abstract

The Bunyamwera (BUNV) orthobunyavirus NSs protein has proven a challenge to study in the context of viral infection. NSs is encoded in a reading frame that overlaps that of the viral nucleocapsid (N) protein thus limiting options for mutagenesis. In addition, NSs is poorly immunogenic, and antibodies only work in certain techniques while the protein itself is subject to proteasomal degradation. In order to generate a virus that expresses NSs independently of N, an ambisense S RNA segment was designed by mutating the 5'- and 3'-terminal nucleotide sequences. These mutations were previously shown to alter promoter activity so that both replication and transcription were promoted from both the genome and the antigenome RNAs (J. N. Barr et al., J. Virol. 79: 12602-12607, 2005). As proof of principle, a recombinant BUNV was created that expressed green fluorescent protein (GFP) in the ambisense orientation. GFP expression was detected throughout at least 10 passages. Recombinant BUNV encoding epitope-tagged versions of NSs in the ambisense orientation expressed NSs via a subgenomic mRNA, and two viruses grew to titers only modestly lower than parental rBUNdelNSs2 virus. The ambisense viruses were temperature sensitive, and NSs was s...Continue Reading

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Citations

Sep 1, 2015·The Journal of General Virology·Benjamin BrennanJohn McLauchlan
Jul 15, 2015·Journal of Virology·Akiko Takenaka-UemaTaisuke Horimoto
May 1, 2018·Frontiers in Veterinary Science·M Fausta DutuzeRebecca C Christofferson

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