| Summary: | Hepatic steatosis is a risk factor for both liver disease progression and an impaired response to interferon alpha (IFN-α)-based combination therapy in chronic hepatitis C virus (HCV) infection. Previously, we reported that free fatty acid (FFA)-treated HCV cell culture induces hepatocellular steatosis and impairs the expression of interferon alpha receptor-1 (IFNAR1), which is why the antiviral activity of IFN-α against HCV is impaired.To investigate the molecular mechanism by which IFNAR1 expression is impaired in HCV cell culture with or without free fatty acid-treatment.HCV-infected Huh 7.5 cells were cultured with or without a mixture of saturated (palmitate) and unsaturated (oleate) long-chain free fatty acids (FFA). Intracytoplasmic fat accumulation in HCV-infected culture was visualized by oil red staining. Clearance of HCV in FFA cell culture treated with type I IFN (IFN-α) and Type III IFN (IFN-λ) was determined by Renilla luciferase activity, and the expression of HCV core was determined by immunostaining. Activation of Jak-Stat signaling in the FFA-treated HCV culture by IFN-α alone and IFN-λ alone was examined by Western blot analysis and confocal microscopy. Lysosomal degradation of IFNAR1 by chaperone-mediated autophagy (CMA) in the FFA-treated HCV cell culture model was investigated.FFA treatment induced dose-dependent hepatocellular steatosis and lipid droplet accumulation in HCV-infected Huh-7.5 cells. FFA treatment of infected culture increased HCV replication in a concentration-dependent manner. Intracellular lipid accumulation led to reduced Stat phosphorylation and nuclear translocation, causing an impaired IFN-α antiviral response and HCV clearance. Type III IFN (IFN-λ), which binds to a separate receptor, induces Stat phosphorylation, and nuclear translocation as well as antiviral clearance in FFA-treated HCV cell culture. We show here that the HCV-induced autophagy response is increased in FFA-treated cell culture. Pharmacological inhibitors of lysosomal degradation, such as ammonium chloride and bafilomycin, prevented IFNAR1 degradation in FFA-treated HCV cell culture. Activators of chaperone-mediated autophagy, including 6-aminonicotinamide and nutrient starvation, decreased IFNAR1 levels in Huh-7.5 cells. Co-immunoprecipitation, colocalization and siRNA knockdown experiments revealed that IFNAR1 but not IFNLR1 interacts with HSC70 and LAMP2A, which are core components of chaperone-mediated autophagy (CMA).Our study presents evidence indicating that chaperone-mediated autophagy targets IFNAR1 degradation in the lysosome in FFA-treated HCV cell culture. These results provide a mechanism for why HCV induced autophagy response selectively degrades type I but not the type III IFNAR1.
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