Fig. 5.

TNFAIP8 suppressed the p53 pathway in NSCLC cells. a NCI-H460 cells infected with lentivirus expressing either Ctrl-shRNA (blue) or TNFAIP8-shRNA2 (red). Genes and samples are listed in the rows and columns, respectively. A colour scale for the normalized expression data is shown at the bottom of the microarray heat map (blue represents downregulated genes; red represents upregulated genes). b Statistically significant modulation (indicated by the inverse log 10 of the P values) of Canonical Pathway following TNFAIP8 knockdown predicted by the commercially available IPA software. c, d qRT-PCR and western blot analyses of p53 and RAD51 expression levels in NCI-H460 and A549 cells treated with TNFAIP8 shRNAs. *P < 0.05 and n.s., not significant (Student’s t-test.) P values and n.s., not significant were calculated using Student’s t-test. e NCI-H460 and A549 cells infected with lentivirus encoding the indicated shRNA were treated with MG132 for 6 h. Lysates were immunoprecipitated with anti-p53 antibody. The ubiquitination of the p53 was analysed by western blotting using anti-ubiquitinantibody. f DNA repair after exposure to cisplatin was shown. A549/cDDP cells were transfected with control shRNA (Ctrl) or TNFAIP8 shRNA2 (TNFAIP8-sh2). Transfected cells were treated with 100 μM cisplatin for 48 h, and RAD51 foci were examined. Scale bar = 5 μM. g, h A549/cDDP cells were transfected with control shRNA (Ctrl) or TNFAIP8 shRNA2 (TNFAIP8-sh2) before cisplatin exposure. Cell mRNA and lysates were prepared after cisplatin exposure, and real-time qRT-PCR and western blotting analyses were performed. i A549/cDDP cells transfected with the indicated constructs were treated with MG132 for 6 h after cisplatin exposure. The ubiquitination of p53 was analysed as above. All n = 3; bar, SEM; n.s., no significant difference; *P < 0.05 (Student’s t-test)