Viral RNA stability and rates of replication in wild-type and C3U variants in the presence of miR-122.

(A) Effect of the C3U mutation on HCV RNA decay. Huh7.5 cells were transfected with wild-type and C3U H77.S3/GLuc RNA, treated with 25 μM of MK-0608 three days later, and viral RNA abundances were monitored at specific times after treatment, using Northern blot analyses. (B) One phase decay graph of HCV RNA (R2 = 0.939–0.947) was determined by normalizing HCV RNA levels to loading control actin from three independent experiments. Estimated half-lives (t1/2) of wild-type and C3U RNA are 3.12 hours and 2.6 hours, respectively. (C) HCV RNA half-lives (t1/2) of three independent experiments (**p<0.0018). (D) Rates of RNA replication in wild-type and mutant HCV variant. Three days after HCV RNA transfection, RNA was labeled with 200 μM EU for 4 and 7 hours, conjugated to biotin and subsequently isolated with streptavidin beads. RNA replication rates were determined by RT-qPCR and data is normalized to wild-type HCV four hour labeling time point (**p<0.0047, ns = non-significant).