Fig. 7: TAK1 siRNA blocks the H5N1 virus-induced degradation of junction proteins.

a A549 cells were transfected with a scrambled control or TAK1 siRNA. After incubation for 24 h, the cells were left uninfected or infected with H5N1 viruses (1 MOI) and incubated for another 24 h. Cell lysates were analyzed for TAK1, ERK, and JNK phosphorylation and for the levels of epithelial junction proteins by Western blot with the indicated antibodies. Relative protein levels were analyzed by quantification of the density of the protein bands with NIH Image-J software and presented as bar graphs. Data are the mean ± SD of three experiments. *p < 0.05, **p < 0.01. b The conditioned media from H5N1 virus-infected cells were analyzed for virus titers by measuring the TCID50 values. c H5N1 virus-induced occludin ubiquitination is blocked by TAK1 or p38 inhibitors. A549 cells were transiently transfected with the expression vector encoding the HA-ubiquitin gene. After incubation for 24 h, the cells were left uninfected or infected with H5N1 viruses (1 MOI) and then incubated in the absence or presence of 5Z (5 μM) or SB202190 (10 μM). Cell lysates were prepared and immunoprecipitated with an antibody against occludin. Whole-cell lysates (WCL) and immunoprecipitates were analyzed with antibodies against ubiquitin, NP, occludin, and β-actin. The data represent one of three independent experiments.