Fig. 2.

The C terminus of NS4B interacts with NS5A in a mammalian two-hybrid system. A. Huh7 cells were transfected with the indicated plasmid combinations. In this assay the interaction between full length NS5A and the C terminal domain (CTD) of NS4B (a.a. 192–261) was examined (see scheme). The culture media was collected 48 h post-transfection and tested for SEAP activity as a control for transfection efficiency. Followed by a luciferase activity assay performed on cell lysates. Luciferase activity results are presented as a percentage relative to the NS5A and NS4B CTD value. Values are (mean±SD) from triplicate wells. The graph is a representative result from three independent experiments. Asterisks denote statistical significance in an unpaired Student t test ^*P=0.01; ^**P=0.008. B. Comparable expression levels of the constructs are shown in panel B as determined by Western blot using VP16 and GAL4 antibodies. Monoclonal mouse anti-β-actin antibody was used as a loading control. Molecular mass markers are indicated on the right.