Hepatocystin/80K-H inhibits replication of hepatitis B virus through interaction with HBx protein in hepatoma cell

Shin, Gu Choul; Ahn, Seung Ho; Choi, Hyosun; Lim, Keo-Heun; Choi, Do Young; Kim, Kwang Pyo; Kim, Kyun-Hwan

doi:10.1016/j.bbadis.2013.04.026

Cited by 14 publications

(13 citation statements)

References 58 publications

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“…Interestingly, comparison of Western blot analysis the clone 13-13 surface antigens (HBsAgs) revealed a completely different band pattern compared to that of WT and clone 13-19, which also harbors the rtA181T mutation. WT HBV expresses three HBsAgs with six bands owing to alternative protein glycosylation, as shown in a previous study (25). However, clone 13-13 showed only one band corresponding to a lower molecular weight than the intact small HBsAg, suggesting the expression of a truncated HBsAgs (Fig.…”

Section: Resultsmentioning

confidence: 91%

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The Impact of the Hepatitis B Virus Polymerase rtA181T Mutation on Replication and Drug Resistance Is Potentially Affected by Overlapping Changes in Surface Gene

Ahn

Park

et al. 2014

J Virol

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The emergence of drug-resistant hepatitis B virus (HBV) is a major problem for antiviral treatment in chronic hepatitis B infection. In this study, we analyzed the evolution of drug-resistant mutations and characterized the effects of the rtA181T and rtI233V mutations on viral replication and drug resistance. We performed a clonal analysis of the HBV polymerase gene from serum samples during viral breakthrough treated with antiviral agents. A series of mutant clones containing rtA181T and/or rtI233V mutations were constructed and determined the effect of these mutations on the replication ability and drug resistance. An in vitro study revealed that the effect of the rtA181T mutation on viral replication and drug resistance is dependent on the mutations in the overlapping surface gene. Compared to the rtA181T surface missense mutation (rtA181T/sW172S), the introduction of rtA181T surface nonsense mutation (rtA181T/sW172*) resulted in decreased viral replication and increased drug resistance. Complementation assay revealed that the truncated PreS1 is responsible for reduced replication of rtA181T/sW172* mutant. Moreover, the rtA181T/sW172* mutant exhibited a defect in viral particle secretion. The rtI233V mutation that emerged during adefovir therapy reduced viral replication and conferred resistance to adefovir. Our data suggest that the impact of the rtA181T mutation on replication and drug resistance differs based on the mutation status of the corresponding surface gene. The rtI233V mutation also affects replication ability and drug resistance. This observation suggests the need for genotypic analysis of overlapping surface genes to manage antiviral drug resistance if clinical isolates harbor the rtA181T mutation. IMPORTANCEThe emergence of drug-resistant HBV that are no longer susceptible to nucleos(t)ide analogues is a major problem for antiviral treatment in chronic hepatitis B infection. Among drug-resistant mutations, the single rtA181T mutation is known to confer cross-resistance to antiviral drugs. This mutation causes intermediate or reduced susceptibility to tenofovir. Moreover, the clinical occurrence of the rtA181T mutation during antiviral therapy is also high. Our study revealed that the effect of the rtA181T mutation on viral replication and drug resistance is dependent on the mutations in the overlapping surface gene. This observation suggests the need for genotypic analysis of overlapping surface genes to manage antiviral drug resistance if clinical isolates harbor the rtA181T mutation. We believe that our study will not only extend the understanding of the drug resistance mechanism, but it will also ultimately provide new treatment options for patients with multidrug resistant HBV.

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Section: Resultsmentioning

confidence: 91%

“…In vitro drug susceptibility and replication assays were performed using Southern blotting as described previously (24,25). Briefly, cell pellets were lysed with 100 l of HEPES buffer containing 1% NP-40.…”

Section: Patientmentioning

confidence: 99%

The Impact of the Hepatitis B Virus Polymerase rtA181T Mutation on Replication and Drug Resistance Is Potentially Affected by Overlapping Changes in Surface Gene

Ahn

Park

et al. 2014

J Virol

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“…HBV genome replication and viral RNAs were detected by Southern and Northern blot analysis, respectively. Southern blot was performed as described previously . Cells were harvested by scrapping at 72 h post‐transfection and lysed with HEPES buffer (10 mmol/L HEPES pH 7.5, 100 mmol/L NaCl, 1 mmol/L EDTA, and 1% NP‐40).…”

Section: Methodsmentioning

confidence: 99%

“…Southern blot was performed as described previously. 20 Cells were harvested by scrapping at 72 h posttransfection and lysed with HEPES buffer (10 mmol/L HEPES pH 7.5, 100 mmol/L NaCl, 1 mmol/L EDTA, and 1% NP-40). Lysate was treated for 15 min with DNase I (1.5 U; Clontech/ Takara Bio, Mountain View, CA, USA) and mung bean nuclease (7.5 U; Clontech/Takara Bio) at 37°C in reaction buffer (500 mmol/L CaCl2 and 800 mmol/L MgCl2) to eliminate transfected plasmid DNA.…”

Section: Quantification Of Hbsagmentioning

confidence: 99%

Pre‐S mutations of hepatitis B virus affect genome replication and expression of surface antigens

Kim

Choi

Ahn

et al. 2014

J of Gastro and Hepatol

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“…Immunoprecipitation and immunoblotting were performed with anti-HBx antibody as described previously. 52 The expression level of TNFSF10 receptors on the cell surface was determined by flow cytometry as described previously. 53 Briefly, cells were detached with diluted trypsin, washed with PBS (ThermoFisher Scientific, 21300025), and resuspended in PBS containing 2% fetal bovine serum.…”

Section: Analysis Of the Expression Of Tnfsf10 Receptor And Hbxmentioning

confidence: 99%

Hepatitis B virus–triggered autophagy targets TNFRSF10B/death receptor 5 for degradation to limit TNFSF10/TRAIL response

et al. 2016

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Death receptors of TNFSF10/TRAIL (tumor necrosis factor superfamily member 10) contribute to immune surveillance against virus-infected or transformed cells by promoting apoptosis. Many viruses evade antiviral immunity by modulating TNFSF10 receptor signaling, leading to persistent infection. Here, we report that hepatitis B virus (HBV) X protein (HBx) restricts TNFSF10 receptor signaling via macroautophagy/autophagymediated degradation of TNFRSF10B/DR5, a TNFSF10 death receptor, and thus permits survival of virusinfected cells. We demonstrate that the expression of the TNFRSF10B protein is dramatically reduced both in liver tissues of chronic hepatitis B patients and in cell lines transfected with HBV or HBx. HBx-mediated downregulation of TNFRSF10B is caused by the lysosomal, but not proteasomal, degradation pathway. Immunoblotting analysis of LC3B and SQSTM1, and microscopy analysis of tandem-fluorescence-tagged LC3B revealed that HBx promotes complete autophagy. Inhibition of autophagy with a pharmacological inhibitor and LC3B knockdown revealed that HBx-induced autophagy is crucial for TNFRSF10B degradation. Immunoprecipitation and GST affinity isolation assays showed that HBx directly interacts with TNFRSF10B and recruits it to phagophores, the precursors to autophagosomes. We confirmed that autophagy activation is related to the downregulation of the TNFRSF10B protein in liver tissues of chronic hepatitis B patients. Inhibition of autophagy enhanced the susceptibility of HBx-infected hepatocytes to TNFSF10. These results identify the dual function of HBx in TNFRSF10B degradation: HBx plays a role as an autophagy receptor-like molecule, which promotes the association of TNFRSF10B with LC3B; HBx is also an autophagy inducer. Our data suggest a molecular mechanism for HBV evasion from TNFSF10-mediated antiviral immunity, which may contribute to chronic HBV infection.

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Hepatocystin/80K-H inhibits replication of hepatitis B virus through interaction with HBx protein in hepatoma cell

Cited by 14 publications

References 58 publications

The Impact of the Hepatitis B Virus Polymerase rtA181T Mutation on Replication and Drug Resistance Is Potentially Affected by Overlapping Changes in Surface Gene

The Impact of the Hepatitis B Virus Polymerase rtA181T Mutation on Replication and Drug Resistance Is Potentially Affected by Overlapping Changes in Surface Gene

Pre‐S mutations of hepatitis B virus affect genome replication and expression of surface antigens

Hepatitis B virus–triggered autophagy targets TNFRSF10B/death receptor 5 for degradation to limit TNFSF10/TRAIL response

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