FIG. 6.

The NES contained in the influenza B virus NEP can be functionally replaced by the influenza A virus NEP NES. (A) Diagram of alterations of the influenza B virus NEP gene. B/delNES/NEP is the wild-type NEP with the first 20 aa removed. A MET codon has been added for proper protein translational initiation. B/NEP is the wild-type NEP gene from influenza B/Lee/40 virus. A21/B/NEP has the first 20 aa of the influenza B virus NEP replaced with the established NES contained within the first 21 aa of the influenza A virus NEP. Both ORFs were cloned into the vector pCAGGS. (B through D) 293T cells were transfected with each of the DNA combinations listed in Table 1. Forty-eight hours posttransfection supernatants were mixed with either 10⁶ PFU of influenza B/Yamagata virus or an equivalent volume of PBS and then used to infect fresh MDCK cells. (B) Primary transfection refers to the CAT activity (10³ dilution of cell extract) from the transfection of 293T cells. (C) Passage+Virus refers to the CAT activity (undiluted cell extract) detected following infection of MDCK cells with supernatants from the 293T transfection mixed with influenza B/Yamagata/73 virus. (D) Passage+PBS is the CAT activity from MDCK cells infected with supernatants from 293T transfected cells and mixed with PBS.