Fig. 3.

Molecular requirements for FnCas9-mediated HCV inhibition. (A) Huh-7.5 cells were transfected with FnCas9 ± NLS, the 5′ UTR-targeting rgRNA, and HCV. At 72 h, viral luciferase was quantified and the percent inhibition compared with the nontargeting rgRNA is displayed (n = 8; data compiled from three independent experiments). (B) Experiments were performed as above, using alanine point mutants in the RuvC domain (D11A), HNH domain (H969A), the double mutant (D11A/H969A), or the ARM (R59A) (n = 8; data compiled from three independent experiments). (C) Rabbit reticulocyte lysate in vitro translation assays of HCV luciferase were performed using the indicated Cas9 and RNAs and viral luciferase measured (n = 4; data are representative of four experiments).