Figure 5.

ART-27 enhances steroid and thyroid receptor-dependent transcriptional activation. HeLa cells (1.2 × 10⁵ cells/35-mm dish) were transiently transfected using Lipofectamine with paired receptor (0.2 μg/dish) and reporter constructs (0.1 μg/dish) for hAR + MMTV-Luc (A), GR + MMTV-Luc (B), ERα and ERβ + XETL (C), and human TRβ-1 + pGL3-DR4 (D) along with the indicated amount of ART-27 or empty expression vector to equalize the total amount of DNA per dish and 0.05 μg of pCMV-LacZ per dish as an internal control for transfection efficiency. Cells were treated with 100 nM R1881 (A), 100 nM dexamethasone (Dex; B), 10 nM 17β-estradiol (E2; C), 100 nM triac (T3, D), or the ethanol vehicle (white bars) for 12 h, and receptor transcriptional activation was assayed as described in MATERIALS AND METHODS, normalized to β-galactosidase activity, and expressed as relative luminescence units (RLU). The average of three independent experiments and SE is shown. (E) ART-27 does not affect transactivation by VP16. HeLa cells were transfected as above with an expression construct for GAL4-VP16 and a reporter plasmid containing five Gal4-binding sites upstream of the E1b promoter, in the absence (white bars) or presence (gray bars) of 1 μg of ART-27, and assayed for luciferase activity. (F) ART-27 failed to activate transcription when tethered to DNA. HeLa cells were transfected as above with expression constructs for LexA, LexA–ART-27, or LexA-AR_18–500 and the pΔ4X-LALO reporter plasmid and assayed for luciferase activity.