Persistent Hepatitis C Virus Infection Impairs Ribavirin Antiviral Activity through Clathrin-Mediated Trafficking of Equilibrative Nucleoside Transporter 1

Panigrahi, Rajesh; Chandra, Partha K.; Ferraris, Pauline; Kurt, Ramazan; Song, Kyoungsub; Garry, Robert F.; Reiss, Krzysztof; Coe, Imogen R.; Furihata, Tomomi; Balart, Luis A.; Wu, Tong; Dash, Srikanta

doi:10.1128/jvi.02492-14

Cited by 21 publications

(18 citation statements)

References 36 publications

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“…Therefore, we speculate that any exogenous agent that induces ER-stress and autophagy response additively inhibits the expression of IFNAR1, leading to impairment of IFN-α treatment response against HCV. Chronic ER-stress and autophagy response also affect RBV antiviral activity by downregulating membrane expression of ENT1 [ 41 ]. Our results provide a mechanism for how FFA-treated HCV cell culture impairs the antiviral response of IFN-α.…”

Section: Discussionmentioning

confidence: 99%

Chaperone-Mediated Autophagy Targets IFNAR1 for Lysosomal Degradation in Free Fatty Acid Treated HCV Cell Culture

et al. 2015

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BackgroundHepatic 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.AimTo investigate the molecular mechanism by which IFNAR1 expression is impaired in HCV cell culture with or without free fatty acid-treatment.MethodHCV-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.ResultsFFA 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).ConclusionOur 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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Section: Discussionmentioning

confidence: 99%

Chaperone-Mediated Autophagy Targets IFNAR1 for Lysosomal Degradation in Free Fatty Acid Treated HCV Cell Culture

et al. 2015

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“…Over the past several years, many investigators, including our own, showed that HCV‐associated ER stress induced an autophagy response, which results in impaired host innate immunity through blocking endogenous IFN production7, 9 and also escapes from exogenously added IFN‐α and ribavirin (RBV)‐based antiviral therapy through degradation of interferon‐alpha receptor 1 and RBV transporter 10, 11. In this report, we found that HCV infection induces chaperone‐mediated autophagy (CMA) as a cell survival mechanism to avoid the ER‐stress response.…”

Section: Introductionmentioning

confidence: 99%

Interferon‐alpha‐induced hepatitis C virus clearance restores p53 tumor suppressor more than direct‐acting antivirals

Aydın

Chatterjee

Chandra

et al. 2017

Hepatology Communications

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The mechanism why hepatitis C virus (HCV) clearance by direct‐acting antivirals (DAAs) does not eliminate the risk of hepatocellular carcinoma (HCC) among patients with advanced cirrhosis is unclear. Many viral and bacterial infections degrade p53 in favor of cell survival to adapt an endoplasmic reticulum (ER)‐stress response. In this study, we examined whether HCV clearance by interferon‐alpha or DAAs normalizes the ER stress and restores the expression of p53 tumor suppressor in cell culture. We found that HCV infection induces chronic ER stress and unfolded protein response in untransformed primary human hepatocytes. The unfolded protein response induces chaperone‐mediated autophagy (CMA) in infected primary human hepatocytes and Huh‐7.5 cells that results in degradation of p53 and induced expression of mouse double minute 2 (Mdm2). Inhibition of p53/Mdm2 interactions by small molecule (nutlin‐3) or silencing Mdm2 did not rescue the p53 degradation, indicating that HCV infection induces degradation of p53 independent of the Mdm2 pathway. Interestingly, we found that HCV infection degrades p53 in a lysosome‐dependent mechanism because lysosome‐associated membrane protein 2A silencing restored p53 degradation. Our results show that HCV clearance induced by interferon‐alpha‐based antiviral therapies normalizes the ER‐stress response and restores p53, whereas HCV clearance by DAAs does neither. We show that decreased expression of p53 in HCV‐infected cirrhotic liver is associated with expression of chaperones associated with ER stress and the CMA response. Conclusion: HCV‐induced ER stress and CMA promote p53 degradation in advanced liver cirrhosis. HCV clearance by DAAs does not restore p53, which provides a potential explanation for why a viral cure by DAAs does not eliminate the HCC risk among patients with advanced liver disease. We propose that resolving the ER‐stress response is an alternative approach to reducing HCC risk among patients with cirrhosis after viral cure. (Hepatology Communications 2017;1:256‐269)

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“…Decreased level of p62 was observed in CHC contrary to AIH, suggestive of an autophagic function occurring at a much higher rate. This is supported by the observation that the level of p62 decreased within 3-6 days following HCV infection in Huh7.5 cells due to an increase in the autophagy response, which was indicated by an elevated level of LC3 [34].…”

Section: Discussionmentioning

confidence: 58%

Autophagy, Mitophagy and MicroRNA Expression in Chronic Hepatitis C and Autoimmune Hepatitis

Szekerczés

Gógl

Illyés

et al. 2020

Pathol. Oncol. Res.

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Although the role of autophagy has been implicated in several forms of chronic hepatitis, it is still not fully understood. Active autophagy eliminates damaged molecules and organelles (such as mitochondria) by lysosomal degradation. In the present study, we aimed to examine and compare autophagy activity in chronic hepatitis C (CHC) and autoimmune hepatitis (AIH) by detecting the expression of autophagy (LC3 and p62) and mitochondrium-related (TOMM20) proteins, as well as the levels of selected microRNAs (miR-101, -155, -204 and − 224) known to be involved in the regulation of autophagy. In addition, the expression levels were related to pathohistological parameters. Liver biopsy samples, including 45 CHC and 18 AIH cases, were immunohistochemically stained for LC3, p62 and TOMM20 and the expression of miRNAs was determined using real-time PCR. We found elevated LC3 and p62 in AIH samples as compared with CHC ones, indicating an activated autophagy that is impaired in AIH as no degradation of p62 seemed to occur. Moreover, p62 showed strong correlation with necroinflammatory grades in the AIH group. The observed elevated levels of TOMM20 and p62 suggest a less efficient elimination of damaged mitochondria in AIH as opposed to CHC, in which autophagy seems to have a more active function. The level of miR-101 was increased in case of CHC as compared with AIH, however, miR-155, -204 and 224 resulted in no expressional. Furthermore, miR-224 level correlated with steatosis and miR-155 expression with fibrosis stage in CHC. In conclusion, dissimilar autophagic activity was observed in CHC and AIH, suggesting a close association between impaired autophagy and severity of necroinflammation. This impairment may not be regulated by the analyzed miRNAs. Nevertheless, miR-224 and − 155 seem to be associated with CHC progression.

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Persistent Hepatitis C Virus Infection Impairs Ribavirin Antiviral Activity through Clathrin-Mediated Trafficking of Equilibrative Nucleoside Transporter 1

Cited by 21 publications

References 36 publications

Chaperone-Mediated Autophagy Targets IFNAR1 for Lysosomal Degradation in Free Fatty Acid Treated HCV Cell Culture

Chaperone-Mediated Autophagy Targets IFNAR1 for Lysosomal Degradation in Free Fatty Acid Treated HCV Cell Culture

Interferon‐alpha‐induced hepatitis C virus clearance restores p53 tumor suppressor more than direct‐acting antivirals

Autophagy, Mitophagy and MicroRNA Expression in Chronic Hepatitis C and Autoimmune Hepatitis

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