Skip to main content
. Author manuscript; available in PMC: 2009 Oct 22.
Published in final edited form as: Cell Microbiol. 2008 Apr 28;10(9):1787–1800. doi: 10.1111/j.1462-5822.2008.01165.x

Figure 3. Expression of IGTP promotes translocation and activity of nuclear transcription factor NF-κB (p65).

Figure 3

A. Tet-On/IGTP HeLa cells were induced with 1 μg/ml Dox, whole cell lysates and nuclear lysates were prepared at indicated time points. NF-κB and I-κBα were detected by Western blot. B. Luciferase assay of IGTP-induced NF-κB activity. (a) Schematic structure of the pNF-κB-Luc plasmid. LUC: firefly luciferase gene; κB4: four copies of the NF-κB consensus enhancer sequence fused to a TATA-like promoter. (b) Luciferase activity assay. Tet-On/IGTP HeLa cells were cotransfected with pNF-κB-luc and pCMV-β-gal control vector, induced with Dox and harvested at indicated time points. Control cells were not induced but cultured for 48 h. Assay reaction and data normalization were performed as described in Experimental Procedures. Data from three independent experiments are presented as a fold increase. Values are means ± SE. *p<0.05. C. Expression of IGTP increases sequence-specific DNA-binding activity of NF-κB. Tet-On/IGTP HeLa cells were induced with Dox and the nuclear proteins were prepared at indicated time points. A gel mobility shift assay was performed using nuclear proteins and 32P-end-labeled oligonucleotides containing a consensus NF-κB binding site.