Helicases DDX5 and DDX17 promote heterogeneity in HBV transcription termination in infected human hepatocytes

• Published in: Journal of hepatology Vol. 81; no. 4; pp. 609 - 620
• Main Authors: Chapus, Fleur, Giraud, Guillaume, Huchon, Pélagie, Rodà, Mélanie, Grand, Xavier, Charre, Caroline, Goldsmith, Chloé, Roca Suarez, Armando Andres, Martinez, Maria-Guadalupe, Fresquet, Judith, Diederichs, Audrey, Locatelli, Maëlle, Polvèche, Hélène, Scholtès, Caroline, Chemin, Isabelle, Hernandez Vargas, Hector, Rivoire, Michel, Bourgeois, Cyril F., Zoulim, Fabien, Testoni, Barbara
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.10.2024; Elsevier
• Subjects: DEAD-box RNA Helicases - genetics; DEAD-box RNA Helicases - metabolism; Gene Expression Regulation, Viral; HBV; Hepatitis B virus - genetics; Hepatitis B virus - physiology; Hepatitis B, Chronic - genetics; Hepatitis B, Chronic - metabolism; Hepatitis B, Chronic - virology; Hepatocytes - metabolism; Hepatocytes - virology; Humans; Life Sciences; Polyadenylation; RNA helicases; RNA polyadenylation; RNA stability; RNA, Viral - genetics; Trans-Activators - genetics; Trans-Activators - metabolism; transcription termination; Transcription Termination, Genetic; Virus Replication - genetics; RNA stability; RNA polyadenylation; RNA helicases; HBV; transcription termination
• ISSN: 0168-8278; 1600-0641; 1600-0641
• DOI: 10.1016/j.jhep.2024.05.016

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. 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. 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. Our findings identify DDX5 and DDX17 as crucial determinants of HBV transcriptional fidelity and as host restriction factors for HBV replication. 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. [Display omitted] •We uncovered a heterogeneous transcriptional termination of HBV transcripts in cellular models and in patients with CHB.•HBV transcription termination is functionally linked to HBV replication.•RNA helicases DDX5 and DDX17 promote HBV transcriptional read-through and viral RNA destabilisation.•DDX5 and DDX17 are host HBV restriction factors and are downregulated in patients with CHB.