Figure 3
The role for NF-κB signaling pathway in regulating transcriptional activity of human Mc1-1 promoter in various human esophageal squamous cell carcinoma cell lines. (A) TE-1, KYSE150 or Eca109 cells were transfected of pGL2-Basic, pMcl-1-κBwt or pMcl-1-κBmt together with pcDNA3.1 empty vector or pcDNA3-DNMIκBα plasmid. The pRL-SV40 was co-transfected in all experiments to correct the variations in transfection efficiency. Transfected cells were incubated for 24 h and then treated with DMSO or 20 μM Bay11-7082 for an additional 12 h after which the activities of firefly and Renilla luciferase were monitored by dual luciferase reporter assay. The relative luciferase activity normalized to the value of Renilla luciferase activity. The Mcl-1 promoter activities were expressed as fold induction of their respective pGL2-Basic-transfected-cells treated with DMSO. Data are shown as means ± S.D. of at least two independent experiments performed in triplicate. Statistical significance: * p < 0.05, compared with their respective pMcl-1-κBwt-transfected-cells treated with DMSO. ** p < 0.05, pMcl-1-κBwt-transfected-cells treated with DMSO versus pMcl-1-κBwt-transfected-cells treated with Bay11-7082. NS, no significant difference between pMcl-1-κBmt-transfected-cells treated with DMSO and pMcl-1-κBmt-transfected-cells treated with Bay11-7082. (B) Effects of Bay11-7082 on the transactivation activity of NF-κB in TE-1 and KYSE150 cell lines. TE-1 or KYSE150 cells were transient transfected with pGL3-Basic vector or pGL3-NF-κB-Luc plasmid and luciferase reporter assay were performed as described in Methods. The relative luciferase activity normalized to the value of Renilla luciferase activity. NF-κB-driven luciferase activities were expressed as fold induction of their respective pGL3-Basic-transfected-cells treated with DMSO. Data are presented as the mean ± S.D. of two independent experiments performed in triplicate. Statistical significance: *, p < 0.01.