Figure 4
NS2B and NS4A promote the degradation of p62. A HEK293T cells were transfected with pCAGGS empty vector, pCAGGS-NS2B-HA, or pCAGGS-NS4A-HA for 24, 36, 48, and 60 h. Samples were harvested for Western blotting analysis and immunoblotted for the proteins p62, LC3, and β-actin. B Normalized p62/β-actin and LC3-II/β-actin intensity band ratios from the data in A. C HEK293T cells were co-transfected with pCAGGS-Flag-p62 and pCAGGS empty vector, pCAGGS-NS2B-HA, or pCAGGS-NS4A-HA for 36 h in a dose dependent manner (0.6, 0.9, 1.2, 1.5 µg). Samples were harvested for Western blotting analysis and immunoblotted for the proteins Flag, HA, and β-actin. D Normalized Flag-p62/β-actin intensity band ratio from the data in C. E HEK293T cells were co-transfected with pCAGGS-Flag-p62 and pCAGGS empty vector, pCAGGS-NS2B-HA, or pCAGGS-NS4A-HA for 36 h. Samples were harvested for Western blotting analysis and coimmunoprecipitation. Anti-Flag antibody which recognizes Flag-p62 was used to coimmunoprecipitate NS2B-HA and NS4A-HA. Student’s t test was performed to determine statistical significance (*, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P lt; 0.0001).