FIG. 6.

RIG-I is responsible for the activation of IRF-3 and production of IFN-α/β in oligodendrocytes induced by MHV infection. (A) Knockdown of RIG-I by siRNAs. N20.1 cells were transfected with a pool of siRNAs specific for RIG-I (RIG-I-siRNA) or the nonspecific control siRNA (CsiRNA). At 48 h posttransfection, intracellular RNAs were isolated and the mRNAs for RIG-I were detected by RT-PCR using primers specific to the mouse RIG-I gene. The housekeeping gene GAPDH was used as an internal control. (B) MHV-induced IRF-3 activation was blocked by knockdown of RIG-I. N20.1 cells were transfected with the RIG-I siRNAs or CsiRNA or mock transfected. At 24 h posttransfection, cells were infected with MHV-A59/GFP at an MOI of 10 or mock infected. At 8 h p.i., nuclear extracts were isolated for determining the IRF-3 activity as described in the legend to Fig. 4. (C and D) Viral recognition by RIG-I is critical for the IFN-α/β induction. Cells were treated as described for panel B. At 8 h p.i. culture medium was collected for determination of IFN-α/β proteins by ELISA. The results are expressed as the mean picograms per ml for three independent experiments. Error bars indicate standard deviations of the means.