tetano
Editor, Senior Moderator
J Gen Virol. 2013 Nov 12. doi: 10.1099/vir.0.056036-0. [Epub ahead of print]
Adaptive mutation in NEP allows stable transgene expression in a chimeric influenza A virus vector.
Kuznetsova I, Shurygina AP, Wolf B, Wolschek M, Enzmann F, Sansyzbay A, Khairullin B, Sandybayev N, Stukova M, Kiselev O, Egorov A, Bergmann M.
Source
Medical University of Vienna;
Abstract
The development of influenza virus vectors with long insertions of foreign sequences remains difficult due to a small size and instable nature of the virus. Here, we used the influenza virus inherent property of self-optimization to generate a vector stably expressing long transgenes from the NS1 open reading frame (NS1-ORF). This was achieved by continuous selection of bright fluorescent plaques of a green fluorescence protein (GFP) expressing vector during multiple passages in mouse B16f1 cells. The newly generated vector acquired stability in interferon (IFN) competent cell lines and in vivo in murine lungs. Although, improved vector fitness was associated with the appearance of 4 coding mutations in PB2, HA and NS segments, the stability of the transgene expression was primarily dependent on the single mutation Q20R in the nuclear export protein (NEP). Importantly, a longer insert, such as a cassette of 1299 nts encoding two Mycobacterium tuberculosis Esat6 and Ag85A proteins, could substitute the GFP-transgene. Thus, the inherent property of the influenza virus to adapt can also be used to adjust a vector backbone to a stable expression of long transgenes.
KEYWORDS:
influenza, vaccine, viral vectors
PMID:
24222196
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24222196
Adaptive mutation in NEP allows stable transgene expression in a chimeric influenza A virus vector.
Kuznetsova I, Shurygina AP, Wolf B, Wolschek M, Enzmann F, Sansyzbay A, Khairullin B, Sandybayev N, Stukova M, Kiselev O, Egorov A, Bergmann M.
Source
Medical University of Vienna;
Abstract
The development of influenza virus vectors with long insertions of foreign sequences remains difficult due to a small size and instable nature of the virus. Here, we used the influenza virus inherent property of self-optimization to generate a vector stably expressing long transgenes from the NS1 open reading frame (NS1-ORF). This was achieved by continuous selection of bright fluorescent plaques of a green fluorescence protein (GFP) expressing vector during multiple passages in mouse B16f1 cells. The newly generated vector acquired stability in interferon (IFN) competent cell lines and in vivo in murine lungs. Although, improved vector fitness was associated with the appearance of 4 coding mutations in PB2, HA and NS segments, the stability of the transgene expression was primarily dependent on the single mutation Q20R in the nuclear export protein (NEP). Importantly, a longer insert, such as a cassette of 1299 nts encoding two Mycobacterium tuberculosis Esat6 and Ag85A proteins, could substitute the GFP-transgene. Thus, the inherent property of the influenza virus to adapt can also be used to adjust a vector backbone to a stable expression of long transgenes.
KEYWORDS:
influenza, vaccine, viral vectors
PMID:
24222196
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24222196