FIG. 5.

Effect of the NEP protein on the ability to package and passage a functional viral RNA-like CAT gene. 293T cells were transfected with either wild-type (WT) or NEP-lacking (−NEP) combinations of plasmids (Table 1). Forty-eight hours posttransfection clarified supernatants were used to infect MDCK cells. Cells were either superinfected with influenza B/Yamagata/73 virus at a multiplicity of 1 or mock infected with PBS. Twelve hours postinfection cells were harvested and assayed for CAT activity. The WT lane shows a positive signal for passage of the viral RNA-like CAT gene. The supernatant derived from the −NEP-transfected cells are not able to passage the viral RNA-like CAT gene. (A) CAT activity (10³ dilution of cell extract) from the primary transfection; (B) CAT activity (undiluted cell extract) detected following infection with clarified supernatants shown in panel A that were superinfected with influenza B/Yamagata/73 virus; (C) CAT activity (undiluted cell extract) from MDCK cells infected with clarified supernatants shown in panel A or mock superinfected with PBS.