35, 36 To

confirm viperin’s anti-HCV activity, we knocked down viperin expression, using an RNA interference (RNAi) approach, and were able to demonstrate, for the first time, Depsipeptide that viperin plays an important, but not exclusive, role in the anti-HCV activity of IFN-α. Considering that many genes are differentially regulated in Huh-7 cells after IFN-α stimulation, it is highly likely that a coordinated ISG response is responsible for the control of HCV replication. A number of studies have suggested that viperin has an ER distribution18, 23; however, we and others have observed that viperin localizes to both LDs and, in our studies, the HCV NS5A-positive RCs.24 LDs have recently been shown to be an essential component of the HCV life cycle,25 and it is thought that the

close association of the LD and ER membranes provides a microenvironment essential for HCV RNA replication and virion production. It has been hypothesized that the interaction of viperin with NS5A at the LD surface is the possible mechanism whereby viperin exerts its antiviral effect through the disruption of virion assembly.12, 13 However, a number of lines of evidence suggest that this is unlikely. First, viperin exerts its anti-HCV effect against the HCV subgenomic replicon, which lacks the HCV structural proteins and is defective in virion assembly. This would also suggest click here that the viperin-core acetylcholine interaction we observed is not fundamental to viperin antiviral activity, and that the interaction with NS5A is critical. It is plausible that the observed interaction between viperin and core at the surface of the LD is mediated by the ability of core to recruit and interact with NS5A at the LD surface to initiate virion assembly. Second,

viperin is antiviral against a genotype 2a HCV subgenomic replicon (SGRm-JFH1BlaRL), in which the HCV IRES drives the expression of the luciferase reporter gene to allow for the quantitative measurement of HCV RNA replication kinetics uncoupled from virion assembly after transfection of in vitro transcribed HCV RNA.10 Expression of viperin significantly suppressed luciferase output from this HCV subgenomic replicon, suggesting that the anti-HCV effect of viperin was at the level of HCV replication and not virion assembly. Finally, through confocal miscroscopy and FRET analysis, we have conclusively shown that viperin interacts with both NS5A and the proviral host factor, VAP-A, within the HCV RC. VAP-A (also known as hVAP-33) is a known interacting partner with NS5A (and NS5B) and is required for the efficient replication of HCV genomic RNA.