Abstract
The number of amino acid substitutions in the interferon (IFN) sensitivity-determining region (ISDR) of hepatitis C virus (HCV) NS5A is a strong predictor for the outcome of IFN-based treatment. To assess the involvement of ISDR in the HCV life cycle and to clarify the molecular mechanisms influencing IFN susceptibility, we used recombinant JFH-1 viruses with NS5A of the genotype 1b Con1 strain (JFH1/5ACon1) and with NS5A ISDR containing 7 amino acid substitutions (JFH1/5ACon1/i-7mut), and compared the virus propagation and the induction of interferon-stimulated genes (ISGs). By transfecting RNAs of these strains into HuH-7-derived cells, we found that the efficiency of infectious virus production of JFH1/5ACon1/i-7mut was attenuated compared with JFH1/5ACon1. After transfecting full-length HCV RNA into HepaRG cells, the mRNA expression of ISGs was sufficiently induced by IFN treatment in JFH1/5ACon1/i-7mut-transfected but not in JFH1/5ACon1-transfected cells. These data suggested that the NS5A-mediated inhibition of ISG induction was deteriorated by amino acid substitutions in the ISDR. In conclusion, using recombinant JFH-1 viruses, we demonstrated that HCV NS5A is associated with infectious virus production and the inhibition of IFN signaling, and amino acid substitutions in the NS5A ISDR deteriorate these functions. These observations explain the strain-specific evasion of IFN signaling by HCV.
Highlights
Hepatitis C virus (HCV) infection is a major cause of chronic liver diseases worldwide
After transfection of full-length hepatitis C virus (HCV) RNA, the replication level of HCV can be assessable by measuring the intracellular HCV core Ag levels, and the efficiency of infectious virus production can be estimable by evaluating the intracellular infectivity titers
The intra- and extracellular infectivity titers of JFH1/5ACon1/i-7mut were significantly lower than those of JFH1/5ACon1 and JFH1/5ACon1/i-wt (Figure 2C). These data suggested that the replication level of the viruses was comparable but that the production of infectious JFH1/5ACon1/i-7mut virus was attenuated compared with the JFH1/5ACon1 and JFH1/5ACon1/i-wt viruses, and this lower virus production was associated with a lower core Ag level in the culture medium of JFH1/5ACon1/i-7mut-transfected cells
Summary
Hepatitis C virus (HCV) infection is a major cause of chronic liver diseases worldwide. HCV establishes a persistent infection and leads to cirrhosis and hepatocellular carcinoma [1, 2]. To eradicate HCV, interferon (IFN)-based treatments have been used for the past two decades; the treatment efficacies have not been satisfactory [3, 4]. The inhibition of immune responses such as attenuation of the IFN-signaling cascade by this virus is thought to be responsible for this insufficient efficacy of IFN-based treatments. Several treatment-refractory cases have been reported, and, in such cases, the evasion strategies against immune system are concerned. It is still important to evaluate the molecular mechanisms that responsible for IFN resistance to understand the strategy the virus uses to establish a persistent infection
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