Fig. 5. NPM overexpression enhances MnSOD gene transcription.
HepG2 cells were co-transfected with either NPM expression vector (pcDNA3.1/NPM) or empty vector (pcDNA3.1) along with Mn-SOD promoter- and enhancer-driven luciferase reporter vector. Parallel transfection experiments were performed using MnSOD promoter- and enhancer-driven luciferase reporter vectors, where the NF-κB binding site in the enhancer region is mutated. After 12 h of co-transfection, cells were washed and grown for 24 h, and then cell lysates were collected. A, equal amounts of cellular proteins were subjected to 10% SDS-polyacrylamide gel electrophoresis, transferred onto nitrocellulose membrane, and then Western blot analyses were conducted with an NPM-V5-epitope antibody. B and C, luciferase activity of cell lysates was measured as a determinant of MnSOD gene transcription. Each data point represents the mean of three independent experiments ± S.D. Significantly different from corresponding empty vector transfected control; **, p < 0.01. IP, immunoprecipitation; WB, Western blot.