Figure 1.
Identification of candidate Foxo-binding sites in the Bim promoters. (A) Map showing four Foxo-binding sites within the Bim promoters (numbered according to the predicted start site reported by Bouillet et al.[15]). (B) EMSA to test for binding of Foxo3a to the Foxo-binding sites in the Bim promoters. 32P-labelled oligos corresponding to the four sites were incubated with nuclear extracts from 293T cells transiently transfected with a control plasmid or tamoxifen-inducible active HA-tagged Foxo3a (HA-Foxo3A(A3)ER) that had been treated for 24 h with tamoxifen. (C) Control and HA-Foxo3a(A3)ER nuclear extracts from (B) were left untreated or incubated with either HA- or Foxo3A-specific antibodies to verify the specificity of Foxo3A binding to site 4 in the Bim promoters. Arrows indicate specific bands in (B) and (C). EMSA, electrophoretic mobility shift assay; FBS, fetal bovine serum; UTR, untranslated region; wt, wild type.