Figure 3

USF 2A inhibits cAMP-mediated induction of RIIβ. A reporter construct containing the region (-395 to -123) from the RIIβ promoter was transfected into Sertoli cell primary cultures together with expression vectors for USF1, USF2a and USF2b isoforms or the corresponding empty expression vector. Empty expression vector was added to ensure a total of 2 μg of DNA transfected in each well. The cells were stimulated for 28 h with 100 μM of 8-CPT-cAMP (black and grey bars) or left untreated (open bars). Reporter activity is shown relative to the cAMP-stimulated level that is set to 100 (black bar). Data are normalized for expression of luciferase from the internal control plasmid. Panel A: Reporter expression levels in unstimulated (open bar) and stimulated cells in the absence (black bar) or presence of expression vectors for USF1 (1), USF2a (2a) or USF2b (2b)(grey bars). Panel B: Reporter levels in the absence (black bar) or presence of different concentrations of USF2a expression vector (0 to 750 ng, grey bars). Three separate transfections were performed in triplicate.