IFN-α inhibits HBV transcription and replication in cell culture and in humanized mice by targeting the epigenetic regulation of the nuclear cccDNA minichromosome
Laura Belloni, … , Maura Dandri, Massimo Levrero
Laura Belloni, … , Maura Dandri, Massimo Levrero
Published February 1, 2012; First published January 17, 2012
Citation Information: J Clin Invest. 2012;122(2):529-537. https://doi.org/10.1172/JCI58847.
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Research Article Virology

IFN-α inhibits HBV transcription and replication in cell culture and in humanized mice by targeting the epigenetic regulation of the nuclear cccDNA minichromosome

Abstract

HBV infection remains a leading cause of death worldwide. IFN-α inhibits viral replication in vitro and in vivo, and pegylated IFN-α is a commonly administered treatment for individuals infected with HBV. The HBV genome contains a typical IFN-stimulated response element (ISRE), but the molecular mechanisms by which IFN-α suppresses HBV replication have not been established in relevant experimental systems. Here, we show that IFN-α inhibits HBV replication by decreasing the transcription of pregenomic RNA (pgRNA) and subgenomic RNA from the HBV covalently closed circular DNA (cccDNA) minichromosome, both in cultured cells in which HBV is replicating and in mice whose livers have been repopulated with human hepatocytes and infected with HBV. Administration of IFN-α resulted in cccDNA-bound histone hypoacetylation as well as active recruitment to the cccDNA of transcriptional corepressors. IFN-α treatment also reduced binding of the STAT1 and STAT2 transcription factors to active cccDNA. The inhibitory activity of IFN-α was linked to the IRSE, as IRSE-mutant HBV transcribed less pgRNA and could not be repressed by IFN-α treatment. Our results identify a molecular mechanism whereby IFN-α mediates epigenetic repression of HBV cccDNA transcriptional activity, which may assist in the development of novel effective therapeutics.

Authors

Laura Belloni, Lena Allweiss, Francesca Guerrieri, Natalia Pediconi, Tassilo Volz, Teresa Pollicino, Joerg Petersen, Giovanni Raimondo, Maura Dandri, Massimo Levrero

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Figure 4

HBV ISRE mediates IFN-α transcriptional repression.

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HBV ISRE mediates IFN-α transcriptional repression. (A) Sequence of the ...
(A) Sequence of the HBV enhancer 1/X gene promoter around the HBV ISRE. ISRE mutations are shown. The nucleotide substitutions do not alter the HBV polymerase polypeptide sequence. (B) Chromatin prepared from untreated and IFN-α–treated HepG2 cells transfected with WT or ISREmt HBV genomes was immunoprecipitated with a relevant control IgG or specific anti-STAT2 antibodies. Immunoprecipitated chromatin was analyzed by qPCR and results expressed as in Figure 1D. (C) mRNAs were prepared from untreated and IFN-α–treated HepG2 cells transfected with WT and ISREmt HBV genomes, and HBV pregenomic RNA was quantified by qPCR using specific primers. GAPDH amplification was used to normalize for equal loading of each RNA sample. (D) Cytoplasmic HBV core particles were isolated from untreated and IFN-α–treated HepG2 cells 48 hours after transfection with monomeric linear full-length WT or ISREmt genomes. Results are expressed as in Figure 1A. Results are shown as mean values from 3 independent experiments; bars indicate SD. P values were determined using Student’s t test. **P < 0.01.