FIGURE 7.

CREB is important for IL-23p19 transcriptional activity. A, DC2.4 cells were transfected with the p19-luciferase construct (p19-GL3), pRL-TK construct, and constructs containing mutations within the distal CREB binding site, the proximal CREB binding site, or both CREB binding sites of p19-GL3. Six hours later the cells were stimulated with TNFα±PGE₁OH for 14 h. Luciferase units were divided by their renilla control, and the empty vector control was then subtracted. -Fold change was calculated by dividing each experimental value by the value for untreated (medium) cells. Reporter activity is presented as -fold change. B and C, DC were treated with TNFα in the presence or absence of PGE₁OH for 2 h. Cells were fixed, sonicated, and subjected to ChIP analysis using antibodies for CREB (black bar) or control GST (white bar). Precipitated DNA was isolated and evaluated by PCR using specific primers for the proximal (−563) and distal (−1125) CREB binding site. **, p < 0.01 and ***, p < 0.001, compared with TNFα (B) and to p19-GL3 (A). N.S., not significant. Data are representative of four independent experiments.