Figure 3.

Role of c-Rel in augmentation of CHGA promoter Hap2 activity. A, pictorial representation of the consensus motif for c-Rel binding as predicted by ConSite. The matrix predicts preferred binding of the transcription factor with the T allele (indicated by star symbol) at both −1018 and −57 bp positions over others. B, representation of the alleles present at the respective SNP loci for the CHGA promoter Hap2 and Hap5. C, effect of overexpression of c-Rel on CHGA Hap2 promoter activity. IMR-32 cells were transfected with CHGA Hap2–GLuc or Hap5–GLuc constructs with increasing doses of c-Rel expression plasmid. pCDNA 3.1(+) was used as balancing plasmid. Results are expressed as mean ± S.E. of triplicate values of the ratio of Gaussia luciferase/μg of protein. **, p < 0.01 as compared with basal activity of Hap2–GLuc construct. Overexpression of c-Rel was confirmed by Western blotting. D, effect of down-regulation of endogenous c-Rel on CHGA Hap2 promoter activity. IMR-32 cells were transfected with CHGA Hap2–GLuc or Hap5–GLuc constructs along with control siRNA oligo or c-Rel siRNA oligo. Results are expressed as mean ± S.E. of triplicate values of the ratio of Gaussia luciferase/μg of protein. ****, p < 0.0001 and ###, p < 0.001 as compared with basal activity of Hap2–GLuc construct. Down-regulation of c-Rel was confirmed by Western blot analysis (shown in the bottom panel).