Hepatitis B Virus X protein is essential to initiate and maintain virus replication after infection

Institute of Virology, Technische Universität München/Helmholtz Zentrum München, Trogerstrasse, 30, 81675 Munich, Germany.
Journal of Hepatology (Impact Factor: 11.34). 03/2011; 55(5):996-1003. DOI: 10.1016/j.jhep.2011.02.015
Source: PubMed


The molecular biology of hepatitis B virus (HBV) has been extensively studied but the exact role of the hepatitis B X protein (HBx) in the context of natural HBV infections remains unknown.
Primary human hepatocytes and differentiated HepaRG cells allowing conditional trans complementation of HBx were infected with wild type (HBV(wt)) or HBx deficient (HBV(x-)) HBV particles and establishment of HBV replication was followed.
We observed that cells inoculated with HBx-deficient HBV particles (HBV(x-)) did not lead to productive HBV infection contrary to cells inoculated with wild type HBV particles (HBV(wt)). Although equal amounts of nuclear covalently closed circular HBV-DNA (cccDNA) demonstrated comparable uptake and nuclear import, active transcription was only observed from HBV(wt) genomes. Trans-complementation of HBx was able to rescue transcription from the HBV(x-) genome and led to antigen and virion secretion, even weeks after infection. Constant expression of HBx was necessary to maintain HBV antigen expression and replication. Finally, we demonstrated that HBx is not packaged into virions during assembly but is expressed after infection within the new host cell to allow epigenetic control of HBV transcription from cccDNA.
Our results demonstrate that HBx is required to initiate and maintain HBV replication and highlight HBx as the key regulator during the natural infection process.

21 Reads
    • "Thus, cccDNA is persistent throughout the cycle of infectivity. cccDNA is decorated with the usual proteins of euchromatin with the addition of two HBV proteins: X and Cp (Belloni et al., 2009; Bock et al., 2001; Buendia and Neuveut, 2015; Ducroux et al., 2014; Fallot et al., 2012; Levrero et al., 2009; Lucifora et al., 2011; Pollicino et al., 2006). "
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatitis B Virus (HBV) is a small virus whose genome has only four open reading frames. We argue that the simplicity of the virion correlates with a complexity of functions for viral proteins. We focus on the HBV core protein (Cp), a small (183 residue) protein that self-assembles to form the viral capsid. However, its functions are a little more complicated than that. In an infected cell Cp modulates every step of the viral lifecycle. Cp is bound to nuclear viral DNA and affects its epigenetics. Cp correlates with RNA specificity. Cp assembles specifically on a reverse transcriptase-viral RNA complex or, apparently, nothing at all. Indeed Cp has been one of the model systems for investigation of virus self-assembly. Cp participates in regulation of reverse transcription. Cp signals completion of reverse transcription to support virus secretion. Cp carries both nuclear localization signals and HBV surface antigen (HBsAg) binding sites; both of these functions appear to be regulated by contents of the capsid. Cp can be targeted by antivirals - while self-assembly is the most accessible of Cp activities, we argue that it makes sense to engage the broader spectrum of Cp function. This article forms part of a symposium in Antiviral Research on "From the discovery of the Australia antigen to the development of new curative therapies for hepatitis B: an unfinished story." Copyright © 2015. Published by Elsevier B.V.
    Antiviral research 06/2015; 121. DOI:10.1016/j.antiviral.2015.06.020 · 3.94 Impact Factor
  • Source
    • "Recent study also showed that the truncation of X protein may result in the inhibition of Core protein although the level of cccDNA didn't change by sgRNA targeting HBx protein (Seeger and Sohn, 2014). It is possible because HBx takes an important role in transcription of preC/C RNA from episomal cccDNA templates (Belloni et al., 2009; Lucifora et al., 2011). Considering the highly economical use of HBV genetic material, whether the inhibition effects caused by frame shift of direct mutagenesis or truncated HBx transcriptional inaction need to be further investigated. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Chronic hepatitis B virus (HBV) infection causes livercirrhosis and hepatocellular carcinoma and remains a serious health problem worldwide. Covalently closed circular DNA (cccDNA) in the liver cell nucleus sustains HBV infection. Major treatments for HBV infection include the use of interferon-α and nucleotide analogs, but they cannot eradicate cccDNA. As a novel tool for genome editing, clustered regularly interspaced short palindromic repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) system developed from bacteria can be used to accurately and efficiently engineer and modify genomic DNA. In this study, the CRISPR/Cas9 system was used to target the HBV genome and efficiently inhibit HBV infection. We synthesized four single-guide RNAs (sgRNAs) targeting the conserved regions of HBV. The expression of these sgRNAS with Cas9 reduced the viral production in Huh7 cells as well as in HBV-replication cell HepG2.2.15. We further demonstrated that CRISPR/Cas9 direct cleavage and cleavage-mediated mutagenesis occurred in HBV cccDNA of transfected cells. In the new mouse model carrying HBV cccDNA, injection of sgRNA-Cas9 plasmids viarapidtailveinresulted in the low level of cccDNA and HBV protein. In conclusion, the designed CRISPR/Cas9 system can accurately and efficiently target HBV cccDNA and inhibit HBV replication. This system may be used as a novel therapeutic strategy against chronic HBV infection. Copyright © 2015. Published by Elsevier B.V.
    Antiviral research 04/2015; 118. DOI:10.1016/j.antiviral.2015.03.015 · 3.94 Impact Factor
  • Source
    • "The cccDNA also functions as an HBV reservoir responsible for persistent replication, and thus is considered to be a reliable marker for HBV infection [Levrero et al., 2009]. Viral HBx has been shown to be essential for the initiation and maintenance of transcription from HBV cccDNA: HBx stimulates the acetylation of histones associated with cccDNA, and is required for transcription in the context of HBV infections [Lucifora et al., 2011]. "
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatitis B virus (HBV) poses a threat to global public health mainly because of complications of HBV-related chronic liver disease. HBV exhibits a narrow host range, replicating primarily in hepatocytes by a still poorly understood mechanism. For the generation of progeny virions, HBV depends on interactions with specific host factors through its life cycle. Revealing and characterizing these interactions are keys to identifying novel antiviral targets, and to developing specific treatment strategies for HBV patients. In this review, recent insights into the HBV-host interactions, especially on virus entry, intracellular trafficking, genome transcription and replication, budding and release, and even cellular restriction factors were reviewed. J. Med. Virol. © 2014 Wiley Periodicals, Inc.
    Journal of Medical Virology 06/2014; 86(6):925-32. DOI:10.1002/jmv.23916 · 2.35 Impact Factor
Show more