Supplementary Figure 2: The mIFIH1 protective variant ablates innate signaling, while mIFIH1^R promotes protection from West Nile virus.

(a-g) HEK293T cells were transfected with 1 μg of plasmid co-expressing either empty vector (EV) or the indicated mIFIH1 constructs described in Fig. 2. (a-b) Data from four biological replicates showing: (b) Geometric MFI and (c) Percent GFP+ for the histograms shown Fig. 2c. (c-d) IFN B1 mRNA expression in cells transfected with mIFIH1^NR or mIFIH1^P constructs at time points indicated showing: (c) Representative experiment from one of three biological replicates; (d) Mean ratio of 3 biological replicates. Error bars represent ± SEM. Statistical analysis using a two-tail student T test. (e-g) At 15 hours post-transfection with the indicated mIFIH1 constructs, cells were infected with West Nile Virus (WNV; MOI of 5) or left uninfected (mock) and harvested at the indicated time points. (e) Quantitative RT-PCR for IFNB1 mRNA expression with data normalized as in Fig. 2d. (f) WNV, IFIH1 and β-actin protein expression assessed by Western blotting. Whole cell lysates were resolved by SDS-PAGE and immuno-blotted with indicated antibodies. (+) represents plasmid or virus present, (-) represents plasmid or virus absent. (f) Densitometry analysis of WNV protein expression at the 30 hours normalized to β-actin levels and displayed as relative fold change. Representative data from one of three WNV infection experiments are displayed. **p<0.01, ****p<0.0001.