TABLE 1.
Synergistic activation of BLV promoter activity by TaxBLV and inhibitors of DNA methylation
Raji cells were transiently cotransfected using the DEAE-dextran procedure with 500 ng of either the pLTRwt-luc (lines 1–6) or pLTR-mut3CRE-luc (lines 7–12) (both methylated in vitro by the SssI methyltransferase before transfection) and with increasing amounts of pSG-TaxBLV (from 0 to 100 ng of plasmid DNA). To maintain the same amount of transfected DNA and to avoid squelching artifacts, the different amounts of pSG-TaxBLV cotransfected were complemented to 100 ng of DNA by using the pSG5 empty vector. Twenty-two hours posttransfection, cells were mock-treated or treated with either 5-AZAdc (250 μm) or procaine (7.5 mm) for 72 h. Luciferase activities were measured in cell lysates 96 h after transfection and normalized with respect to protein concentrations. Results are presented as relative light units (RLU), TaxBLV fold activation, TaxBLV + 5-AZAdc fold activation, and TaxBLV + procaine fold activation of the reporter constructs (pLTRwt-luc and pLTR-mut3CRE-luc) with regard to their respective basal activity level, which was arbitrarily set at a value of 1. The TaxBLV + 5-AZAdc (or TaxBLV + procaine) fold synergism was determined as described previously (51) using the following formula: [Fold activation by TaxBLV + 5-AZAdc (or TaxBLV + procaine)]/{[fold activation by TaxBLV alone] + [fold activation by 5-AZAdc (or procaine) alone]}. Values represent the means of triplicate samples with S.E. indicated in parentheses. An experiment representative of four independent transfections is shown.
