FIG. 2.

Both c-Jun and c-Fos inhibit T-Ag-mediated JCV DNA replication. A replication-competent plasmid, pBLCAT₃-Mad-1L (5 μg), containing the regulatory region of the Mad-1 strain of JCV was transfected alone or in combination with c-Jun (RSV-c-Jun), c-Fos (RSV-c-Fos), and T-Ag (CMV JCV T-Ag) expression plasmids into U-87MG cells, as described in Materials and Methods. Plasmid DNA concentrations used in transfections are indicated on top (in micrograms per 60-mm-diameter plate). The total amount of DNA transfected into the cells was normalized with appropriate empty vectors. At 72 h posttransfection, low-molecular-weight DNA was isolated by the Hirt method (22), digested with both BamHI and DpnI enzymes, and analyzed by Southern blotting. The bands corresponding to the replicated DNA bands were quantitated by a densitometric method (see Materials and Methods) and expressed as percent inhibition with respect to the viral DNA replication in the presence of T-Ag alone. Representative data for the DpnI assay are shown, and variability among different experimental data is indicated by standard deviation. In lane 1, pBLCAT3-Mad-1L plasmid linearized with BamHI enzyme digestion was loaded as a control. The results are represented as percent inhibition and are shown at the bottom. The variability between different replication assays is indicated by standard deviation.