Enhanced virus translation enables miR-122-independent Hepatitis C Virus propagation

The 5UTR of the Hepatitis C Virus genome forms RNA structures that regulate virus replication and translation. The region contains a viral internal ribosomal entry site and a 5 terminal region. Binding of the liver specific miRNA, miR-122, to two conserved binding sites in the 5 terminal region regulates viral replication, translation, and genome stability, and is essential for efficient virus replication, but its precise mechanism of its action is still under debate. A current hypothesis is that miR-122 binding stimulates viral translation by facilitating the viral 5 UTR to form the translationally active HCV IRES RNA structure. While miR-122 is essential for detectable virus replication in cell culture, several viral variants with 5 UTR mutations exhibit low level replication in the absence of miR-122. We show that HCV mutants capable of replicating independently of miR-122 also replicate independently of other microRNAs generated by the canonical miRNA synthesis pathway. Further, we also show that the mutant genomes display an enhanced translation phenotype that correlates with their ability to replicate independently of miR-122. Finally, we provide evidence that translation regulation is the major role for miR-122, and show that miR-122-independent HCV replication can be rescued to miR-122-dependent levels by the combined impacts of 5 UTR mutations that stimulate translation, and by stabilizing the viral genome by knockdown of host exonucleases and phosphatases that degrade the genome. Thus, we provide a model suggesting that translation stimulation and genome stabilization are the primary roles for miR-122 in the virus life cycle. IMPORTANCEThe unusual role of miR-122 in promoting HCV propagation is incompletely understood but is essential for an HCV infection. To better understand its role, we have analyzed HCV mutants capable of replicating independently of miR-122. Our data show that the ability of viruses to replicate independently of miR-122 correlates with enhanced virus translation, but that genome stabilization is required to restore efficient HCV replication. This suggests that viruses must gain both abilities to escape the need for miR-122 and impacts the possibility that HCV can evolve to replicate outside of the liver.