Biology subjects

Chapus, F.

Publications and source records attributed to Chapus, F..

3 recordsLinked to original sources

bioRxiv2025

DDX5 and DDX17 RNA helicases regulate hepatitis B virus RNA splicing

Chronic HBV infection remains a major health burden worldwide and is the main driver of severe liver diseases. Liver pathogenesis is associated with the increased proportion of HBV spliced variants that encode viral proteins involved in liver disease progression. However, how HBV RNA splicing is regulated is poorly understood. Here, we focused on DDX5 and DDX17 RNA helicases, known to regulate HBV RNA metabolism and alternative splicing of host genes. By performing 5RACE-PCR combined with single molecule sequencing, we demonstrated that silencing both proteins increased the usage of a specific splicing donor site and the expression of the derived HBV spliced variants. Polysome fractionation highlighted the ability of these RNA species to encode new viral proteins potentially contributing to liver pathogenesis. Overall, our data established DDX5 and DDX17 helicases as master regulators of HBV RNA metabolism, by fine-tuning viral splicing, which is linked to HBV-induced liver pathogenesis and disease progression.

molecular biology↗

bioRxiv2024

Helicases DDX5 and DDX17 promote Hepatitis B Virus transcription termination heterogeneity in infected human hepatocytes

Background & AimsTranscription termination fine tunes gene expression and contributes to specify the function of RNAs in eukaryotic cells. Transcription termination of hepatitis B virus (HBV) is subjected to the recognition of the canonical polyadenylation signal (cPAS) common to all viral transcripts. The regulation of the usage of this cPAS and its impact on viral gene expression and replication is currently unknown. Approach & ResultsTo unravel the regulation of HBV transcript termination, we implemented a 3 RACE-PCR assay coupled to single molecule sequencing both in in vitro infected hepatocytes and in chronically infected patients. 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 to HBV RNA destabilization and decreased HBx protein expression. RNA and chromatin immunoprecipitation, together with mutation of cPAS sequence suggested a direct role of DDX5 and DDX17 in functionally linking cPAS recognition to transcriptional readthrough, HBV RNA stability and replication. ConclusionsOur findings identify DDX5 and DDX17 as crucial determinants for HBV transcriptional fidelity and as host restriction factors for HBV replication.

molecular biology↗

bioRxiv2020

The CCCTC-binding factor CTCF represses hepatitis B virus Enhancer I and regulates viral transcription

Hepatitis B virus (HBV) infection is of global importance with over 2 billion people exposed to the virus during their lifetime and at risk of progressive liver disease, cirrhosis and hepatocellular carcinoma. HBV is a member of the hepadnaviridae family that replicates via episomal copies of a covalently closed circular DNA (cccDNA) genome. The chromatinization of this small viral genome, with overlapping open reading frames and regulatory elements, suggests an important role for epigenetic pathways to regulate viral transcription. The chromatin-organising transcriptional insulator protein CCCTC-binding factor (CTCF) has been reported to regulate transcription in a diverse range of viruses. We identified two conserved CTCF binding sites in the HBV genome within Enhancer I and chromatin immunoprecipitation (ChIP) analysis demonstrated an enrichment of CTCF binding to integrated or episomal copies of the viral genome. siRNA knockdown of CTCF results in a significant increase in pre-genomic RNA levels in de novo infected HepG2 cells and those supporting episomal HBV DNA replication. Furthermore, mutation of these sites in HBV DNA minicircles abrogated CTCF binding and increased pre-genomic RNA levels, providing evidence of a direct role for CTCF in repressing HBV transcription. IMPORTANCEHepatitis B virus (HBV) is a global cause of liver disease. At least 300 million individuals are chronically infected with HBV, frequently leading to life-threatening liver cirrhosis and cancer. Following viral entry, HBV DNA enters the nucleus and is bound by histones that are subject to epigenetic modification. The HBV genome contains two enhancer elements that stimulate viral transcription but the interplay between the viral enhancers and promoters is not fully understood. We have identified the host cell protein CCCTC binding factor (CTCF) as a repressor of HBV gene expression. CTCF binds to the HBV genome within Enhancer I and represses transcription of pre-genomic RNA. These findings provide new insights into how HBV transcription is regulated and show a new role for CTCF as a transcriptional insulator by associating with the viral genome between Enhancer I and the downstream basal core promoter.

microbiology↗