Transcriptional activation of RTA on the ORF59 promoters in a dose-dependent manner. The reporter plasmid pGL3-ORF59pFL contains a 1909-bp sequence upstream of the start codon of ORF59 that drives the expression of firefly luciferase. A fixed amount (1 μg) of the reporter plasmids was cotransfected into 10 million 293T (A) and DG75 (B) cells with 0, 0.5, 1, 2.5, 5 and 10 μg of pCR3.1-RTA and 10, 9.5, 9, 7.5, 5 and 0 μg of pCR3.1 control vector. 50 ng pRL-Tk was also included in each transfection, and served as an internal control for transfection efficiency. 24 h posttransfection, cell lysate of each transfection was assayed with the dual-luciferase system. Firefly luciferase activities were normalized to the corresponding Renilla luciferase activities. Fold activation by RTA was calculated by comparing the normalized firefly luciferase activity stimulated by pCR3.1-RTA to that stimulated by pCR3.1. The means and standard deviations from three independent transfections are shown.