Figure 10.

FOXO1 suppression of GnRH-induced Fshb transcription involves two elements in the proximal Fshb promoter. (A) Diagram illustrating the mutations in the AP1 binding element (AP1BE) in the murine Fshb promoter. (B) The −398 Fshb-luc reporter and the Fshb AP1 mutation (mut) were transiently transfected into LβT2 cells along with pcDNA3 empty vector (EV) or FOXO1, as indicated. After overnight incubation in serum-free media, cells were treated for 6 hours with 0.1% BSA vehicle or 10 nM GnRH. The results represent the mean ± SEM of three experiments performed in triplicate and are presented as fold GnRH induction relative to the vehicle control. *, Fshb-luc transcription is significantly repressed by FOXO1 compared to EV using Student's t test; #, the AP1 mut was significantly repressed compared to the wild-type Fshb promoter. (C) The −398 Fshb-luc reporter and 10-bp deletions ranging from −70/−61 to −20/−11 were transiently transfected into LβT2 cells along with EV or FOXO1, as indicated. After overnight incubation in serum-free media, cells were treated for 6 hours with 0.1% BSA or 10 nM GnRH. The results represent the mean ± SEM of three experiments performed in triplicate and are presented as fold GnRH induction relative to the vehicle control. *, Fshb-luc transcription is significantly repressed by FOXO1 compared to EV using one-way ANOVA followed by Tukey's HSD post-hoc test.