Fig. 6.

The miR-33/33*-AMPK axis modulates MAVS activation by interfering with mitophagy. a and b Confocal fluorescence microscopy of HeLa cells transfected with miRNAs, siRNAs or plasmids as indicated, along with Mito-YFP (1 μg/ml) and LC3-mCherry (1 μg/ml) plasmids, followed by infection for 12 h with VSV (MOI = 1). Enlarged area, enlargement of the area outlined on the left; scale bar, 2 µm. c Immunoblot analysis of the indicated proteins in HeLa cells transfected with miRNAs, siRNAs, or plasmids as indicated followed by VSV stimulation for 12 h. d Flow cytometry analysis of HeLa cells transfected with miRNAs, siRNAs, or plasmids as indicated and stained with the mitochondrial superoxide–specific stain MitoSOX after VSV infection for 12 h. e Flow cytometry analysis of HeLa cells transfected as in d and stained with JC-1 during VSV infection for 12 h. f Immunoblot analysis of the indicated proteins of HeLa cells transfected with control miRNAs, siRNAs or vectors, miR-33/33* in AMPKα expressing vectors and MFF siRNA (MFF in) as indicated followed by VSV stimulation for 12 h. g Flow cytometry analysis of HeLa cells transfected as in f stained with the mitochondrial superoxide–specific stain MitoSOX after VSV infection for 12 h. h Flow cytometry analysis of HeLa cells transfected as in f and stained with JC-1 after VSV infection for 12 h. i Macrophages transfected with nc or miR-33/33* were pretreated with MitoTEMPO (0.5–1 μm) as indicated, followed by VSV infection for 8 h (MOI = 1), and MAVS aggregation was analyzed by SDD-AGE and SDS-PAGE. Cells transfected with control miRNAs, siRNAs plus EV, are referred to as control in this figure. Data are representative of three independent experiments.