FIG 3.

Virus-induced IFN-I response and mitochondrial apoptosis are RIG-I dependent. (A) Depletion of RIG-I by RNAi. A549 cells were first reverse transfected with siRNA for RIG-I or control siRNA (C), followed by forward transfection 24 h later. Efficiency of depletion was assessed 72 h after the first transfection by Western blotting using specific antibody against RIG-I. (B) Depletion of RIG-I prevents induction of IFN-I by VSV and SeV. Cells depleted of RIG-I, as described for panel A, were infected with VSV (MOI of 3) and SeV (80 hemagglutinin units/ml) for 16 h or left uninfected (u). Cells were lysed, total RNA was extracted, and cDNA was synthesized by reverse transcription. IFN-β mRNA was detected by RT-qPCR using specific TaqMan probes for IFN-β and GAPDH. Gene expression levels relative to GAPDH were determined according to the 2^−ΔΔCT method. Data represent fold induction relative to levels in uninfected cells (n = 3; means ± standard deviations). (C) Induction of apoptosis by SeV and VSV is dependent on RIG-I. Cells depleted for RIG-I, as described for panel A, were infected with VSV (MOI of 3) or SeV (80 hemagglutinin units/ml). Cleavage of PARP was assessed by Western blotting using a specific antibody. Tubulin expression was used as a loading control.