Fig 5.
Impacts of escape mutations inside the LLRHHNMVY2450-2458 epitope and at covariant sites on HCV replication. The impacts of different mutations were determined in a transient replication assay utilizing the HCV genotype 1b subgenomic replicon Con1. (A) Alignment of the different variants that have been tested. Note that the prototype sequence of Con1 already contains two substitutions compared to the genotype 1b consensus sequence (M2456L and S2439T). (B) Luciferase activities normalized to the 4-h result for each construct over a time course of 96 h. (C) Luciferase activities normalized to the results for the consensus sequence at 48 h (peak of replication) and 96 h (end of experiment). Data in panels B and C are mean values for three independent repetitions, each performed in duplicate, with standard errors of the means. RLU, relative light units. (D) Competition assay between a replicon with the consensus genotype 1b sequence in the epitope region and a replicon with the H2454R substitution. Consensus and variant RNAs were mixed at a ratio of 1:10 before transfection. After 4 h, 7 days, and 18 days, multiple clones of the replicons were sequenced to determine the relative frequencies of the consensus and variant replicons.