Abstract
BackgroundIn hepatocyte nuclei, hepatitis B virus (HBV) genomes occur episomally as covalently closed circular DNA (cccDNA). The HBV X protein (HBx) is required to initiate and maintain HBV replication. The functional nuclear localization of cccDNA and HBx remains unexplored.ResultsTo identify virus–host genome interactions and the underlying nuclear landscape for the first time, we combined circular chromosome conformation capture (4C) with RNA-seq and ChIP-seq. Moreover, we studied HBx-binding to HBV episomes. In HBV-positive HepaRG hepatocytes, we observed preferential association of HBV episomes and HBx with actively transcribed nuclear domains on the host genome correlating in size with constrained topological units of chromatin. Interestingly, HBx alone occupied transcribed chromatin domains. Silencing of native HBx caused reduced episomal HBV stability.ConclusionsAs part of the HBV episome, HBx might stabilize HBV episomal nuclear localization. Our observations may contribute to the understanding of long-term episomal stability and the facilitation of viral persistence. The exact mechanism by which HBx contributes to HBV nuclear persistence warrants further investigations.
Highlights
In hepatocyte nuclei, hepatitis B virus (HBV) genomes occur episomally as covalently closed circular relaxed circular DNA (DNA)
Its virus particles contain a partially double-stranded, relaxed circular DNA genome (RC DNA), and HBV features a complex replication cycle: Upon infection mediated by the Na+-taurocholate cotransporting polypeptide (NTCP) receptor, the HBV nucleocapsids become transported to the hepatocellular nucleus, wherein the RC DNA becomes converted into a circular episome, the covalently closed circular DNA
(early in the HBV life cycle) after nuclear entry, the establishment of HBV closed circular DNA (cccDNA) episomality is based on the conversion of the partially single-stranded HBV RC DNA to the double-stranded cccDNA at a time, when HBV X protein (HBx) is virtually not yet available
Summary
Hepatitis B virus (HBV) genomes occur episomally as covalently closed circular DNA (cccDNA). Chronic HBV infection is estimated to account for more than 80% of HCC cases [4]. Its virus particles contain a partially double-stranded, relaxed circular DNA genome (RC DNA), and HBV features a complex replication cycle: Upon infection mediated by the NTCP receptor, the HBV nucleocapsids become transported to the hepatocellular nucleus, wherein the RC DNA becomes converted into a circular episome, the covalently closed circular DNA (cccDNA). The cccDNA serves as a template for all HBV RNA synthesis and persists in the hepatocyte nucleus [6]. This double-stranded genome is transcribed as a long pregenomic RNA (pgRNA) and subsequently converted to DNA through reverse transcription. Current antiviral therapy is not curative as it predominantly targets cytoplasmic reverse transcription of the pgRNA, but not the episomal persistence
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