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Competing and noncompeting activities of miR-122 and the 5' exonuclease Xrn1 in regulation of hepatitis C virus replication.

ABSTRACT: Hepatitis C virus (HCV) replication is dependent on microRNA 122 (miR-122), a liver-specific microRNA that recruits Argonaute 2 to the 5' end of the viral genome, stabilizing it and slowing its decay both in cell-free reactions and in infected cells. Here we describe the RNA degradation pathways against which miR-122 provides protection. Transfected HCV RNA is degraded by both the 5' exonuclease Xrn1 and 3' exonuclease exosome complex, whereas replicating RNA within infected cells is degraded primarily by Xrn1 with no contribution from the exosome. Consistent with this, sequencing of the 5' and 3' ends of RNA degradation intermediates in infected cells confirmed that 5' decay is the primary pathway for HCV RNA degradation. Xrn1 knockdown enhances HCV replication, indicating that Xrn1 decay and the viral replicase compete to set RNA abundance within infected cells. Xrn1 knockdown and miR-122 supplementation have equal, redundant, and nonadditive effects on the rate of viral RNA decay, indicating that miR-122 protects HCV RNA from 5' decay. Nevertheless, Xrn1 knockdown does not rescue replication of a viral mutant defective in miR-122 binding, indicating that miR-122 has additional yet uncharacterized function(s) in the viral life cycle.

SUBMITTER: Li Y

PROVIDER: S-EPMC3562843 | biostudies-other | 2013 Jan

REPOSITORIES: biostudies-other

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Competing and noncompeting activities of miR-122 and the 5' exonuclease Xrn1 in regulation of hepatitis C virus replication.

Li You Y Masaki Takahiro T Yamane Daisuke D McGivern David R DR Lemon Stanley M SM

Proceedings of the National Academy of Sciences of the United States of America 20121217 5

Hepatitis C virus (HCV) replication is dependent on microRNA 122 (miR-122), a liver-specific microRNA that recruits Argonaute 2 to the 5' end of the viral genome, stabilizing it and slowing its decay both in cell-free reactions and in infected cells. Here we describe the RNA degradation pathways against which miR-122 provides protection. Transfected HCV RNA is degraded by both the 5' exonuclease Xrn1 and 3' exonuclease exosome complex, whereas replicating RNA within infected cells is degraded pr ...[more]

PMID: 23248316

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Project description:The molecular mechanisms of liver pathology and clinical disease in hepatitis E virus (HEV) infection remain unclear. MicroRNAs (miRNAs) are known to modulate viral pathogenesis either by directly altering viral gene expression or by enhancing cellular antiviral responses. Given the importance of microRNA-122 (miR-122) in liver pathobiology, we investigated possible role of miR-122 in HEV infection. In silico predictions using HEV genotype 1 (HEV-1), HEV-2, HEV-3, and HEV-4 sequences showed that the majority of genomes (203/222) harbor at least one miR-122/microRNA-122-3p (miR-122*) target site. Interestingly, HEV-1 genomes showed a highly (97%) conserved miR-122 target site in the RNA-dependent RNA polymerase (RdRp) region (RdRpc). We analyzed the significance of miR-122 target sites in HEV-1/HEV-3 (HEV-1/3) genomes by using a replicon-based cell culture system. HEV infection did not change the basal levels of miR-122 in hepatoma cells. However, transfection of these cells with miR-122 mimics enhanced HEV-1/3 replication and depletion of miR-122 with inhibitors led to suppression of HEV-1/3 replication. Mutant HEV-1 replicons with an altered target RdRpc sequence (CACTCC) showed a drastic decrease in virus replication, whereas introduction of alternative miR-122 target sites in mutant replicons rescued viral replication. There was enrichment of HEV-1 RNA and miR-122 molecules in RNA-induced silencing complexes in HEV-infected cells. Furthermore, pulldown of miR-122 molecules from HEV-infected cells resulted in pulldown of HEV genomic RNA along with miR-122 molecules. These observations indicate that miR-122 facilitates HEV-1 replication, probably via direct interaction with a target site in the viral genome. The positive role of miR-122 in viral replication presents novel opportunities for antiviral therapy and management of hepatitis E.IMPORTANCE Hepatitis E is a problem in both developing and developed countries. HEV infection in most patients follows a self-limited course; however, 20% to 30% mortality is seen in infected pregnant women. HEV superinfections in patients with chronic hepatitis B or hepatitis C virus infections are associated with adverse clinical outcomes, and both conditions warrant therapy. Chronic HEV infections in immunocompromised transplant recipients are known to rapidly progress into cirrhosis. Currently, off-label use of ribavirin (RBV) and polyethylene glycol-interferon (PEG-IFN) as antiviral therapy has shown promising results in both acute and chronic hepatitis E patients; however, the teratogenicity of RBV limits its use during pregnancy, while alpha IFN (IFN-α) increases the risk of transplant rejections. Experimental data determined with genotype 1 virus in the current study show that miR-122 facilitates HEV replication. These observations present novel opportunities for antiviral therapy and management of hepatitis E.

Dataset Information

Competing and noncompeting activities of miR-122 and the 5' exonuclease Xrn1 in regulation of hepatitis C virus replication.

Publications

Competing and noncompeting activities of miR-122 and the 5' exonuclease Xrn1 in regulation of hepatitis C virus replication.

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