Figure 6.

T₄ recruits certain coactivators to TRα1 with potency that was nearly equal to that of T₃ in vitro. Full-length in vitro–transcribed and translated, ³⁵S-labeled human TRα1 or TRβ1 were incubated with glutathione-agarose bead immobilized GST fusions of coactivator proteins (LXXLL receptor interaction motifs) indicated within each panel: SRC1 (amino acids 568–891) (A), TRAP220 (486–723) (B), CBP (1–451) (C), and GRIP (544–767) (D). Incubations were conducted in the presence of the indicated T₃ or T₄ concentration. The immobilized GST constructs were washed, and the nuclear receptors remaining bound to each construct were eluted. The resulting coactivator-TR complexes were characterized by SDS-PAGE and phosphorimager analysis and plotted as the percent maximum binding for each coactivator. Error bars indicate SEs of at least 3 replicate experiments. E, Reverse experiment was conducted with full-length in vitro–transcribed and translated ³⁵S-labeled GRIP1 with immobilized full-length GST-TRα1 or GST-TRβ1, with vehicle alone or the indicated T₃ or T₄ hormone concentration. Vertical dashed and solid lines represent EC50s T3 and T4, respectively.