FIG 6.

The effects of miR-122 binding at each site are cooperative rather than merely multiplicative. Subgenomic miR-122 binding site mutants from Fig. 4B and C were evaluated in Huh7.5 cells for the contribution of miR-122 binding at each site to the overall replication of the viral RNA on day 3. Geometric mean fold increase in luciferase expression of subgenomic S2:p3 and S1:p3 viral RNAs over S1+S2:p3 [Unbound] is shown for two independent experiments and is representative of three independent experiments. S2:p3 [S1 Bound] and S1:p3 [S2 Bound] were the indicated viral RNAs supplemented with miControl. Theoretical [S1+S2 Bound] was determined by multiplying the fold increase of [S1 Bound] by the fold-increase of [S2 Bound] to show the theoretical impact of binding at both sites, if their contribution was multiplicative. Experimental S2:p3 [S1+S2 Bound] and Experimental S1:p3 [S1+S2 Bound] were the indicated viral RNAs supplemented with miR-122/p3. Error bars are geometric standard deviations.