Adaptive mutation in nuclear export protein allows stable transgene expression in a chimaeric influenza A virus vector

作者：Kuznetsova Irina; Shurygina Anna Polina; Wolf Brigitte; Wolschek Markus; Enzmann Florian; Sansyzbay Abylay; Khairullin Berik; Sandybayev Nurlan; Stukova Marina; Kiselev Oleg; Egorov Andrej; Bergmann Michael^*

来源：Journal of General Virology, 2014, 95(2): 337-349.

DOI：10.1099/vir.0.056036-0

The development of influenza virus vectors with long insertions of foreign sequences remains difficult due to the 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 protein ORF. This was achieved by continuous selection of bright fluorescent plaques of a GFP-expressing vector during multiple passages in mouse B16f1 cells. The newly generated vector acquired stability in IFN-competent cell lines and in vivo in murine lungs. Although improved vector fitness was associated with the appearance of four coding mutations in the polymerase (PB2), haemagglutinin and non-structural (NS) segments, the stability of the transgene expression was dependent primarily on the single mutation Q20R in the nuclear export protein (NEP). Importantly, a longer insert, such as a cassette of 1299 nt encoding two Mycobacterium tuberculosis Esat6 and Ag85A proteins, could substitute for the GFP transgene. Thus, the inherent property of the influenza virus to adapt can also be used to adjust a vector backbone to give stable expression of long transgenes.

• 出版日期2014-2