Antiviral Autophagy Pathway Analysis

HEK293 cells, HeLa cells, U2OS cells and Vero cells were cultured in Dulbecco’s modified Eagle medium (Invitrogen, Carlsbad, CA, USA) supplemented with 10% fetal bovine serum (Hyclone, South Logan, UT, USA) and antibiotics (100 U ml⁻¹ penicillin and 100 μg ml⁻¹ streptomycin, Invitrogen). Antibodies used in this study for western blot or immunofluorescence assays were as follows: mouse anti-Flag (Sigma, St Louis, MO, USA), rabbit anti-LC3 (Novus, Littleton, CO, USA and MBL, Woburn, MA, USA), mouse anti-TIM23 (BD, Waltham, MA, USA), mouse anti-GAPDH and rabbit anti-Caspase-3 (Sungene Biotech, Tianjin, China), rabbit anti-Myc and rabbit anti-HA (MBL), rabbit anti-ATG5 (Epitomics, Cambridge, MA, USA), and rabbit anti-TOM20, mouse anti-Calnexin, mouse anti-TRAF2 and rabbit anti-TRAF6 (Santa Cruz Biotechnology, Dallas, TX, USA), rabbit anti-Cleaved Caspase-3 (Cell Signalling Technology, Danvers, MA, USA); the antibody against MAVS was produced by our lab [14 (link)]. Mito-TEMPO was purchased from Santa Cruz Biotechnology or Enzo Life Science, Farmingdale, NY, USA. Pyrrolidinedithiocarbamic acid ammonium salt (PDTC), bafilomycin A1 and chloroquine were purchased from Sigma. VSV was propagated in Vero cells.