FIG. 2.

The LR is required for KSHV Rta to activate transcription. Human endothelial SLK cells were cotransfected with the indicated amounts of vector expressing WT ORF50 or ORF50ΔLR and a luciferase reporter vector for the K-bZIP (A) or Nut-1/PAN (B) promoter. Total DNA was normalized by the addition of empty expression vectors. The cells were harvested and lysed at 40 h posttransfection. Luciferase activity was normalized to β-galactosidase activity, and n-fold activation is given relative to that of the reporter vector transfected alone (0 μg plasmid). Vertical bars indicate standard deviations. Data represent results of triplicate transfection experiments repeated at least twice. See Materials and Methods for details. (C) Equivalent amounts of WT ORF50 and ORF50ΔLR were expressed in transfected SLK cells. The extracts used for the experiments shown in panel A were analyzed by SDS-PAGE/Western blotting, using anti-Rta serum. (D) WT ORF50 and ORF50ΔLR are both expressed in SLK cell nuclei. SLK cells were transfected as for panel A and analyzed by immunofluorescence 40 h posttransfection, using anti-Rta primary serum, fluorescein isothiocyanate-conjugated secondary serum, and DAPI (4′,6′-diamidino-2-phenylindole) to stain nuclear DNA. Vec, vector.