FIG. 4.

Kinase activity of IKK is required for optimum NF-κB activation by SRC-3. (A) HeLa cells were transfected as described in the legend for Fig. 3. The IKK inhibitor, NaSal, was added to a final concentration of 5 mM after transfection and incubated for another 24 h. Thereafter, the luciferase activity was determined and expressed as described earlier. (B) As a control, HeLa cells were transfected with a reporter gene (5× UAS TATA luciferase) along with either GAL4 or GAL4-VP16 expression plasmid, and the results showed that neither the basal (GAL4) (bottom left panel) nor the activated (GAL4-VP16) (bottom right panel) activity of this promoter was affected. RLU, relative light units. SRC-3 and the indicated IKK were cotranslated and labeled with [³⁵S]methionine in vitro. The products were then immunoprecipitated with anti-HA antibody, resolved by SDS-PAGE, and visualized by autoradiography.