Dataset Information

Cell-based cccDNA reporter assay combined with functional genomics identifies YBX1 as HBV cccDNA host factor and antiviral candidate target.

ABSTRACT:

Objectives

Chronic hepatitis B virus (HBV) infection is a leading cause of liver disease and hepatocellular carcinoma. A key feature of HBV replication is the synthesis of the covalently close circular (ccc)DNA, not targeted by current treatments and whose elimination would be crucial for viral cure. To date, little is known about cccDNA formation. One major challenge to address this urgent question is the absence of robust models for the study of cccDNA biology.

Design

We established a cell-based HBV cccDNA reporter assay and performed a loss-of-function screen targeting 239 genes encoding the human DNA damage response machinery.

Results

Overcoming the limitations of current models, the reporter assay enables to quantity cccDNA levels using a robust ELISA as a readout. A loss-of-function screen identified 27 candidate cccDNA host factors, including Y box binding protein 1 (YBX1), a DNA binding protein regulating transcription and translation. Validation studies in authentic infection models revealed a robust decrease in HBV cccDNA levels following silencing, providing proof-of-concept for the importance of YBX1 in the early steps of the HBV life cycle. In patients, YBX1 expression robustly correlates with both HBV load and liver disease progression.

Conclusion

Our cell-based reporter assay enables the discovery of HBV cccDNA host factors including YBX1 and is suitable for the characterisation of cccDNA-related host factors, antiviral targets and compounds.

SUBMITTER: Verrier ER

PROVIDER: S-EPMC10423543 | biostudies-literature | 2022 Dec

REPOSITORIES: biostudies-literature

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Publications

Cell-based cccDNA reporter assay combined with functional genomics identifies YBX1 as HBV cccDNA host factor and antiviral candidate target.

Verrier Eloi R ER Ligat Gaëtan G Heydmann Laura L Doernbrack Katharina K Miller Julija J Maglott-Roth Anne A Jühling Frank F El Saghire Houssein H Heuschkel Margaux J MJ Fujiwara Naoto N Hsieh Sen-Yung SY Hoshida Yujin Y Root David E DE Felli Emanuele E Pessaux Patrick P Mukherji Atish A Mailly Laurent L Schuster Catherine C Brino Laurent L Nassal Michael M Baumert Thomas F TF

Gut 20221209

<h4>Objectives</h4>Chronic hepatitis B virus (HBV) infection is a leading cause of liver disease and hepatocellular carcinoma. A key feature of HBV replication is the synthesis of the covalently close circular (ccc)DNA, not targeted by current treatments and whose elimination would be crucial for viral cure. To date, little is known about cccDNA formation. One major challenge to address this urgent question is the absence of robust models for the study of cccDNA biology.<h4>Design</h4>We establi ...[more]

PMID: 36591611

Similar Datasets

Project description:BackgroundHepatitis B virus covalently closed circular DNA (HBV cccDNA) is assembled by histones and non-histones into a chromatin-like cccDNA minichromosome in the nucleus. The cellular histone acetyltransferase GCN5, displaying succinyltransferase activity, is recruited onto cccDNA to modulate HBV transcription in cells. Clinically, IFN-α is able to repress cccDNA. However, the underlying mechanism of IFN-α in the depression of cccDNA mediated by GCN5 is poorly understood. Here, we explored the effect of IFN-α on GCN5-mediated succinylation in the epigenetic regulation of HBV cccDNA minichromosome.ResultsSuccinylation modification of the cccDNA minichromosome has been observed in HBV-infected human liver-chimeric mice and HBV-expressing cell lines. Moreover, histone H3K79 succinylation by GCN5 was identified in the system. Interestingly, the mutant of histone H3K79 efficiently blocked the replication of HBV, and interference with GCN5 resulted in decreased levels of HBV DNA, HBsAg, and HBeAg in the supernatant from de novo HBV-infected HepaRG cells. Consistently, the levels of histone H3K79 succinylation were significantly elevated in the livers of HBV-infected human liver-chimeric mice. The knockdown or overexpression of GCN5 or the mutant of GCN5 could affect the binding of GCN5 to cccDNA or H3K79 succinylation, leading to a change in cccDNA transcription activity. In addition, Southern blot analysis validated that siGCN5 decreased the levels of cccDNA in the cells, suggesting that GCN5-mediated succinylation of histone H3K79 contributes to the epigenetic regulation of cccDNA minichromosome. Strikingly, IFN-α effectively depressed histone H3K79 succinylation in HBV cccDNA minichromosome in de novo HepG2-NTCP and HBV-infected HepaRG cells.ConclusionsIFN-α epigenetically regulates the HBV cccDNA minichromosome by modulating GCN5-mediated succinylation of histone H3K79 to clear HBV cccDNA. Our findings provide new insights into the mechanism by which IFN-α modulate the epigenetic regulation of HBV cccDNA minichromosome.

Dataset Information

Cell-based cccDNA reporter assay combined with functional genomics identifies YBX1 as HBV cccDNA host factor and antiviral candidate target.

Objectives

Design

Results

Conclusion

Publications

Cell-based cccDNA reporter assay combined with functional genomics identifies YBX1 as HBV cccDNA host factor and antiviral candidate target.

Similar Datasets

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