Fig. 2.

Knockdown of TRIM14 impairs the cellular antiviral response. (A) A549 human lung epithelial cells were transfected with control or TRIM14-specific siRNA oligos as indicated. Forty-eight hours posttransfection, Western blot analysis was used to determine the expression of TRIM14 or actin. (B) A549 cells were transfected with indicated siRNA oligos and an ISRE reporter plasmid. After 48 h, cells were left uninfected or were infected with SeV. Cells were harvested at 12 hpi and analyzed for luciferase activity to determine ISRE activity. Data are mean values ± SD (n = 3). *P < 0.05. (C) A549 cells were transfected with indicated siRNA oligos. After 48 h, cells were left uninfected or were infected with SeV. Cells were harvested at 6 or 12 hpi for Western blot analysis. (D) A549 cells were transfected with indicated siRNA oligos and an NF-κB reporter plasmid. After 48 h, cells left uninfected or infected with SeV for 12 h were analyzed for NF-κB activity. Data are mean values ± SD (n = 3). *P < 0.05. (E) A549 cells were treated as indicated in C and were analyzed by Western blot with indicated antibodies. (F) A549 cells were transfected with indicated siRNA oligos for 48 h. Cells then were transfected with poly(I:C) (500 ng/mL). After 24 h, total RNA was analyzed using real-time quantitative PCR to determine IFNB1 expression. Data are mean values ± SD (n = 3). **P < 0.01. (G) A549 cells were transfected with indicated siRNA oligos for 48 h. Then cells were left uninfected or were infected with SeV. After 16 h, total RNA was analyzed using RT-PCR to determine expression levels of IFNB1, ISG15, TNFA, or GAPDH (control). (H) HEK293T cells were transfected with indicated siRNA oligos. After 48 h, cells were infected with VSV at an MOI of 0.01. Supernatants were harvested at 16 hpi. The virus titers were measured using standard plaque assays. Data are mean values ± SD (n = 3). **P < 0.01.