Additional file 1 of MARCH8 promotes the proteasomal degradation of foot-and-mouth disease virus VP1, VP2, and VP3 to negatively regulate viral replication

Additional file 1. K210R mutant FMDV is not resistant to MARCH8 inhibition. (A) Schematic representation of an FMDV infectious clone. (B) BHK21 cells were infected with rO/HN/93, rO/HN/93-K210R, rO/HN/93-K63R, rO/HN/93-K118R, rO/HN/93-KR, or O/HN/9 for 8 h. The cells were fixed and incubated with primary anti-FMDV serum, followed by staining with FITC-conjugated secondary antibodies (green). Nuclei were stained with DAPI (blue). The fluorescence was observed using a fluorescence inverted microscope. (C and D) BHK21 cells were infected with 0.01 MOI O/HN/93, rO/HN/93, or rO/HN/93-K210R for the indicated time points. The virus titres were determined by a plaque assay. (E and F) SK6 cells in 6-well plates were transfected with 2 μg pCDNA3.1Flag or pCDNA3.1Flag-MARCH8 for 24 h, followed by infection with 0.01 MOI O/HN/93, rO/HN/93, or rO/HN/93-K210R for 8 h. Viral replication was determined by western blotting and plaque assays. The data are shown as the means ± SD. *, P < 0.05; **, P < 0.01; ***, P < 0.001.