Dissecting the Roles of the 5 = Exoribonucleases Xrn1 and Xrn2 in Restricting Hepatitis C Virus Replication

The replication of hepatitis C virus (HCV) is uniquely dependent on a host microRNA, miR-122. Previous studies using genotype 1a H77S.3 virus demonstrated that miR-122 acts in part by protecting the RNA genome from 5 = decay mediated by the cytoplas-mic 5 = exoribonuclease, Xrn1. However, this finding has been challenged by a recent report suggesting that a predominantly nu-clear exoribonuclease, Xrn2, mediates the degradation of genotype 2a JFH1 RNA. Here, we dissect the roles of these two 5 = exori-bonucleases in restricting the replication of different HCV strains andmediating the decay of HCV RNA. Small interfering RNA (siRNA) depletion experiments indicated that Xrn1 restricts replication of all HCV strains tested: JFH1, H77S.3, H77D (a ro-bustly replicating genotype 1a variant), and HJ3-5 (a genotype 1a/2a chimeric virus). In contrast, the antiviral effects of Xrn2 were limited to JFH1 and H77D viruses. Moreover, such effects were not apparent in cells infected with a JFH1 luciferase re-porter virus. Whereas Xrn1 depletion significantly slowed decay of JFH1 and HJ3-5 RNAs, Xrn2 depletion marginally enhanced the JFH1 RNA half-life and had no effect on HJ3-5 RNA decay. The positive effects of Xrn1 depletion on JFH1 replication were largely redundant and nonadditive with those of exogenous miR-122 supplementation, whereas Xrn2 depletion acted additively and thus independently of miR-122. We conclude that Xrn1 is the dominant 5 = exoribonuclease mediating decay of HCV RNA and that miR-122 provides protection against it. The restriction of JFH1 and H77D replication by Xrn2 is likely indirect in na-ture and possibly linked to cytopathic effects of these robustly replicating viruses