Fig. 5: WDR77 impedes MAVS to form prion-like filaments.

a A two-step assay was performed. In step 1, HEK293T cells were infected with SeV for 8 h and harvested to get S5 fraction. Increasing amounts of S5 fractions were incubated with P5 fractions from WT or WDR77^-/- HEK293T cells at 30 °C for 1 h. P5 fractions were then isolated. In step 2, P5 fractions from step 1 were incubated with S5 fractions from WT HEK293T cells at 30 °C for another 1 h. IRF3 dimerization was then analyzed by native PAGE and MAVS aggregation was analyzed by SDD-AGE. Blots shown are representative of n = 2 biological replicates. b A two-step assay was performed. In step 1, S5 fractions were prepared from WT and WDR77^-/- HEK293T cells infected with SeV for 8 h, which were incubated with P5 fractions from WT HEK293T cells at 30 °C for 1 h. P5 fractions were then isolated. In step 2, P5 fractions from step 1 were incubated with S5 fractions from WT HEK293T cells at 30 °C for another 1 h. IRF3 dimerization was analyzed by native PAGE and MAVS aggregation was analyzed by SDD-AGE. Blots shown are representative of n = 2 biological replicates. c Recombinant MAVS was separated into two peaks by gel filtration chromatography, i.e., Peak-I and Peak-II. MAVS (Peak-II) was incubated with BSA or recombinant WDR77 at 4 °C. At various time points, the mixtures were subjected to negative staining and examination under electron microscopy. Scale bars indicate 100 nm. Representative images of n = 3. d MAVS (Peak-I) was incubated with BSA or recombinant WDR77 at 4 °C. At various time points, the mixtures were subjected to negative staining and examination under electron microscopy. Scale bars indicate 100 nm. Data are representative of two independent experiments with similar results (a, b), or three independent experiments with similar results (c, d). Source data are provided as a Source Data file.