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BACKGROUND & AIMS: Transcription termination fine-tunes gene expression and contributes to the specification of RNA function in eukaryotic cells. Transcription termination of HBV is subject to the recognition of the canonical polyadenylation signal (cPAS) common to all viral transcripts. However, the regulation of this cPAS and its impact on viral gene expression and replication is currently unknown. METHODS: To unravel the regulation of HBV transcript termination, we implemented a 3' RACE (rapid amplification of cDNA ends)-PCR assay coupled to single molecule sequencing both in in vitro-infected hepatocytes and in chronically infected patients. RESULTS: The detection of a previously unidentified transcriptional readthrough indicated that the cPAS was not systematically recognized during HBV replication in vitro and in vivo. Gene expression downregulation experiments demonstrated a role for the RNA helicases DDX5 and DDX17 in promoting viral transcriptional readthrough, which was, in turn, associated with HBV RNA destabilization and decreased HBx protein expression. RNA and chromatin immunoprecipitation, together with mutation of the cPAS sequence, suggested a direct role of DDX5 and DDX17 in functionally linking cPAS recognition to transcriptional readthrough, HBV RNA stability and replication. CONCLUSIONS: Our findings identify DDX5 and DDX17 as crucial determinants of HBV transcriptional fidelity and as host restriction factors for HBV replication. IMPACT AND IMPLICATIONS: HBV covalently closed circular (ccc)DNA degradation or functional inactivation remains the holy grail for the achievement of HBV cure. Transcriptional fidelity is a cornerstone in the regulation of gene expression. Here, we demonstrate that two helicases, DDX5 and DDX17, inhibit recognition of the HBV polyadenylation signal and thereby transcriptional termination, thus decreasing HBV RNA stability and acting as restriction factors for efficient cccDNA transcription and viral replication. The observation that DDX5 and DDX17 are downregulated in patients chronically infected with HBV suggests a role for these helicases in HBV persistence in vivo. These results open new perspectives for researchers aiming at identifying new targets to neutralise cccDNA transcription.
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Hepatitis B is a "silent epidemic" that is fifty to a hundred (50-100) times more infectious than HIV and is a potentially life-threatening liver infection [...].
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A sentinel study on viral hepatitis is currently being carried out in the village of Cavunge in a semiarid rural region of the state of Bahia, northeastern Brazil. This study has identified individuals in whom anti-HBc IgG was the only serological marker for hepatitis B virus (HBV). This serological pattern may constitute evidence of occult HBV infection. This study Investigated the possibility of occult hepatitis B virus infection in individuals in a rural community who tested positive for anti-HBc IgG alone. A cross-sectional population-based study. ELISA III was performed on serum samples to test for serological viral markers, and ultrasensitive PCR (US-PCR) was used to assess viremia. Among the 1,536 serum samples, 3.6 percent (n=55) were positive for anti-HBc alone. Four years after this first serological survey, 31 of those 55 individuals (56.3 percent) were retested, and 11 (35.5 percent) remained anti-HBc positive alone. Two of these 31 (6.5 percent) were HBV-DNA positive based on US-PCR, with normal aminotransferase levels in both cases. Cases of occult hepatitis B infection were identified in this semiarid rural community of northeastern Brazil, where endemicity of HBV is moderate.