Hepatitis B virus genome recycling and de novo secondary infection events maintain stable cccDNA levels
| Autor: | Ke Zhang, Karin Wisskirchen, Anindita Chakraborty, Theresa Asen, Wen Min Chou, Wang-Shick Ryu, Jane A. McKeating, Julia Hasreiter, Ulrike Protzer, Romina Bester, Daniela Stadler, Chunkyu Ko, Jochen M. Wettengel |
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| Rok vydání: | 2018 |
| Předmět: |
0301 basic medicine
Hepatitis B virus Secondary infection Organic Anion Transporters Sodium-Dependent Genome Viral Biology medicine.disease_cause Virus Replication Virus Article Polyethylene Glycols 03 medical and health sciences chemistry.chemical_compound 0302 clinical medicine Hbv Hepatitis B Virus Cccdna Ntcp Replenishment Viral Spread Transmission Intracellular Recycling Transcription (biology) medicine Humans Dimethyl Sulfoxide Hepatology Nucleoside analogue Symporters Coinfection cccDNA Hep G2 Cells Hepatitis B Virology 3. Good health HBx 030104 developmental biology chemistry DNA Viral RNA Viral 030211 gastroenterology & hepatology DNA Circular DNA medicine.drug Half-Life |
| Zdroj: | Journal of Hepatology J Hepatol J. Hepatol. 69, 1231-1241 (2018) |
| DOI: | 10.1016/j.jhep.2018.08.012 |
| Popis: | Background & Aims: Several steps in the HBV life cycle remain obscure because of a lack of robust in vitro infection models. These steps include particle entry, formation and maintenance of covalently closed circular (ccc) DNA, kinetics of gene expression and viral transmission routes. This study aimed to investigate infection kinetics and cccDNA dynamics during long-term culture.Methods: We selected a highly permissive HepG2-NTCP-K7 cell clone engineered to express sodium taurocholate cotransporting polypeptide (NTCP) that supports the full HBV life cycle. We characterized the replication kinetics and dynamics of HBV over six weeks of infection.Results: HBV infection kinetics showed a slow infection process. Nuclear cccDNA was only detected 24 h post-infection and increased until 3 days post-infection (dpi). Viral RNAs increased from 3 dpi reaching a plateau at 6 dpi. HBV protein levels followed similar kinetics with HBx levels reaching a plateau first. cccDNA levels modestly increased throughout the 45-day study period with 5-12 copies per infected cell. Newly produced relaxed circular DNA within capsids was reimported into the nucleus and replenished the cccDNA pool. In addition to intracellular recycling of HBV genomes, secondary de novo infection events resulted in cccDNA formation. Inhibition of relaxed circular DNA formation by nucleoside analogue treatment of infected cells enabled us to measure cccDNA dynamics. HBV cccDNA decayed slowly with a half-life of about 40 days.Conclusions: After a slow infection process, HBV maintains a stable cccDNA pool by intracellular recycling of HBV genomes and via secondary infection. Our results provide important insights into the dynamics of HBV infection and support the future design and evaluation of new antiviral agents.Lay summary: Using a unique hepatocellular model system designed to support viral growth, we demonstrate that hepatitis B virus (HBV) has remarkably slow infection kinetics. Establishment of the episomal transcription template and the persistent form of the virus, so called covalently closed circular DNA, as well as viral transcription and protein expression all take a long time. Once established, HBV maintains a stable pool of covalently closed circular DNA via intracellular recycling of HBV genomes and through infection of naive cells by newly formed virions. (C) 2018 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved. |
| Databáze: | OpenAIRE |
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