Figure 6. ATAD1 also augmented the antiviral function of MAVS, independent of its action on NS5B.

• A, B
  Extensive cleavage of MAVS in ATAD^KO cells upon HCV infection. WT, ATAD1^KO, and ATAD1^KO+Res Huh7.5 cells were infected with HCV 2a JFH1 for 48 h. The cells were harvested and lysed with the IP lysis buffer containing phosphatase inhibitor cocktails, and analyzed by western blotting with antibodies anti‐MAVS, anti‐Core, anti‐TBK1, anti‐p‐TBK1, etc. Extensive cleavage of MAVS in ATAD1^KO Huh7.5 cells was restored in ATAD1^KO+Res cells, accompanied by increased phosphorylation level of TBK1.
• C
  ATAD1 promoted the aggregates of MAVS upon HCV infection. WT, ATAD1^KO, and ATAD1^KO+Res Huh7.5 cells were infected with J6/JFH1‐EGFPΔ40 for 72 h, followed by sorting the HCV‐positive cells using FACS. Then, HCV‐infected cells and control cells were transfected with DsRed‐Mito. The cells were fixed and blotted with mouse primary antibody anti‐MAVS for 2 h at room temperature, followed by anti‐mouse IgG (H+L), F(ab′)₂ fragment (Alexa Fluor® 647 Conjugate) for 1 h at room temperature. The images were captured using Nikon Eclipse Ni‐E. The prion‐like aggregates of MAVS (green) and mitochondria (red) were analyzed. Scale bars, 10 μm.
• D
  Knockout of ATAD1 impaired the aggregation of MAVS. WT and ATAD1^KO Huh7.5 cells were infected with JFH1 for 72 h. The cells were harvested and analyzed using SDD‐AGE with anti‐MAVS.
• E
  ATAD1 remained negatively regulating HCV infection without MAVS. WT, ATAD1^KO, MAVS^KO, and MAVS^KO/ATAD1^KO Huh7.5 cells were infected with JFH1 for 48 h. The cells were harvested and analyzed by western blotting with anti‐MAVS and anti‐Core antibodies. An increased level of HCV infection was observed in MAVS^KO/ATAD1^KO cells compared to MAVS^KO cells (right panel).

Source data are available online for this figure.