FoxP3 inhibits the DNA-binding and transcriptional activity of AP-1. (A) HEK 293 cells were transfected without or with FoxP3 and c-Jun expression plasmids as indicated. Nuclear extracts were isolated. Equal amounts of nuclear proteins were incubated with biotin-labeled AP-1 promoter fragments, and gel shift assay was performed (top panel). The protein expression levels of c-Jun in the nuclear extracts were detected by Western blotting using anti-Jun antibody (middle panel). The densities of these AP-1 DNA-binding bands were measured using NIH 1.63 software (bottom panel). Error bars represent data from 3 independent experiments. Data were statistically analyzed with Student t test. (B) AP-1 luciferase, c-Jun, and FoxP3 expression plasmids were cotransfected into HEK 293 cells. The luciferase activities were determined. Error bars represent data from 3 independent experiments. (C) IL-2 luciferase reporter plasmid was cotransfected without or with FoxP3 plasmid into HEK 293 cells. The luciferase activities were determined. Error bars represent data from 3 independent experiments.