Fig. 4

NF-κB binds to the promoter region of GGT1 to activate its transcription. A Protein levels of GGT1 and p65 in MCF-7 and MDA-MB-231 cells transfected with p65 overexpression plasmid (P65-OE) or empty vector were examined by western blot. B RNA level of GGT1 in MCF-7 and MDA-MB-231 cells transfected with p65 overexpression plasmid (P65-OE) or empty vector was examined by qRT-PCR. C Protein levels of GGT1 and p65 in MCF-7 and MDA-MB-231 cells treated with BAY 11–7802 (1 μM) or DMSO were examined by western blot. D RNA level of GGT1 in MCF-7 and MDA-MB-231 cells treated with BAY 11–7802 (1 μM) or DMSO was examined by qRT-PCR. E Schematic of predicted binding sites between GGT1 promoter and p65. F-G Construction of luciferase reporter vectors comprising p65 binding sites in the DNA promoter region of GGT1 (2 kb). Dual-luciferase reporter assays were performed by transfecting the promoter region of GGT1 (GGT1 Pro) or pGL3-basic plasmid in MCF-7 and MDA-MB-231 cells treated with BAY 11–7802 (1 μM) or DMSO. H-I The binding of p65 to the GGT1 promoter region in MCF-7 and MDA-MB-231 cells treated with BAY 11–7802 (1 μM) or DMSO was examined by ChIP assay. Results are shown are shown as mean ± S.D from at least three independent experiments. *p < 0.05; **p < 0.01; ***p < 0.001 (unpaired two-tailed Student’s t test)