FIG. 1.

LCMV Z inhibits CAT expression by the LCMVSCAT2 minigenome in a dose-dependent manner. BHK-21 cells were infected with vTF7.3 (MOI = 3) and subsequently cotransfected with 0.5 μg of pLCMVSCAT2, 1.5 μg of pCITE-NP, 0.1 μg of pGEM-L, 0.1 μg of pTMI-GFP, and increasing amounts of pUCIRES-Z as indicated. Plasmid pGEM-L was not added in lane 8. In all of the samples the total amount of DNA was kept constant (2.7 μg) by adding the appropriate amount of plasmid pTMI. The use of pTMI-GFP allowed us to determine the efficiency of transfection, based on number of GFP-positive cells, prior harvesting of the cells for CAT assays, and protein expression analysis. (A) LCMVSCAT2 minigenome expression. At 24 h after transfection, cell lysates were prepared for measuring CAT activity as described in Material and Methods. (B) Z protein expression. Cell lysates were subjected to SDS-PAGE, and the levels of Z protein expression were determined by Western blot using a rabbit antibody to Z. Lysate applied in lane 1 was prepared from cells infected with LCMV (MOI = 3) at 24 h p.i. The position of the Z protein is indicated on the right, and the molecular size markers are indicated on the left. O, origin; Cm, chloramphenicol; MAc, monoacetylated chloramphenicol; DAc, diacetylated chloramphenicol.