Fig. 1.

CAR3 transactivation of a DR-4 × 3 reporter is ligand-dependent and facilitated by the overexpression of RXR. In A and B, COS-1 cells were transfected with indicated expression vectors [A, pcDNA3.1 (empty vector), B, pcDNA3.1/RXR] in combination with TK-luciferase reporters containing three copies of a direct repeat spaced by 1 to 5 base pairs. A diagram of the vector scheme and details of the respective direct repeat elements used for the experiments conducted in A (−RXR) and B (+ RXR) are shown at the top. Cells were transfected for 18 h as described under Materials and Methods. On day 2, transfected cells were treated with either DMSO or 5 μM CITCO (A and B) or increasing amounts of CITCO as indicated (C) for 24 h, after which luciferase activity was assayed. CMV2 is an empty expression vector. Data are presented as normalized and adjusted luciferase values in which the activity of the DMSO-treated CMV2/3.1 (A), DMSO-treated CMV2/RXR (B), or 500 pM CITCO-treated CMV2/3.1 (C) data point is set to 1. Each data point represents the mean (± S.E.) of four separate transfections.