FIG. 4.

E-box DNA-binding activity increases during Sertoli cell differentiation. In EMSAs, radiolabeled probes containing a consensus E-box that is selective for USF binding (A) or a region of the Fshr promoter containing an E-box known to bind USF proteins (B) were incubated with nuclear extracts isolated from cultured 5-, 11-, and 20-day-old Sertoli cells that were treated with vehicle or T4 (10⁻⁶ M). Representative images of the resulting DNA-protein complexes are shown, and quantitation of the mean (±SEM) of three independent experiments is provided. The relative binding activity for each condition is normalized to that of vehicle-treated 11-day-old Sertoli cells (=1). Values with different lowercase letters differ significantly (P < 0.05). C) USF1 and USF2 are responsible for the increase in DNA-binding activity during Sertoli cell differentiation. DNA-protein complexes from nuclear extracts isolated from cultured 11-day-old Sertoli cells were incubated with nonimmune sera or antisera for USF1, USF2, E47, E2A, or MYC. Supershifted DNA-protein complexes containing USF1 or USF2 are indicated. Exposure times for all complexes are less than 48 h.