FIG. 2.

Cotransfection of Tax with luciferase reporter plasmids in 293 cells. (A) Luciferase assay in which 0.25 μg of pRS-HTax1C was cotransfected with 0.25 μg of the indicated reporter plasmids in 5 × 10⁴ 293 cells with FuGene6 reagent (Roche) in 24-well plates. In experiments using Elk-1, either the Elk-1 expression plasmid (pElk-1-RSPA) or pBS-RSPA was transfected at 0.25 μg per well. The reporter plasmids are shown on the y axis, and activation levels by Tax are represented on the x axis. The experiment was repeated at least three times, and the error bars represent the standard deviations. (B) Expression level of Elk-1 in 293 cells as shown by Western blotting. Lysates are labeled at the top. Lanes 1 and 4 were loaded with mock-transfected lysates, lanes 2 and 5 were loaded with lysates from cells transfected with pElk-1-RSPA, and lanes 3 and 6 were loaded with lysates from cells transfected with both Elk-1 and Tax expression plasmids. Lysates were analyzed on 4 to 12% Bis-Tris gels (Novex), probed with a 1:1,000 dilution of rabbit anti-Elk-1 (αElk-1) (New England Biolabs) and rabbit anti-Tax antibodies, and visualized by chemiluminescence (New England Biolabs).