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Journal article
Characterization of the intracellular deproteinized relaxed circular DNA of hepatitis B virus: An intermediate of covalently closed circular DNA formation
Journal of virology, v 81(22), pp 12472-12484
01 Nov 2007
PMID: 17804499
Featured in Collection : UN Sustainable Development Goals @ Drexel
Abstract
Covalently closed circular DNA (cccDNA) of hepatitis B virus (HBV) is formed by conversion of capsid-associated relaxed circular DNA (rcDNA) via unknown mechanisms and exists in the nucleus of the infected hepatocyte as a minichromosome that serves as the transcription template for viral RNAs. To study the molecular pathway of cccDNA formation and its regulation by viral and cellular factors, we have established a cell line that supports the replication of an envelope protein-deficient HBV genome in a tetracycline-inducible manner. Following induction of HBV replication, the cells accumulate higher levels of cccDNA as well as larger amounts of deproteinized rcDNA (DP-rcDNA) than cells that replicate wild-type HBV genomes. These results indicate that HBV envelope proteins negatively regulate cccDNA formation, and conversion of DP-rcDNA into cccDNA is a rate-limiting step of cccDNA formation in HepG2 cells. Detailed analyses reveal the following: (i) DP-rcDNA exists in both cytoplasm and nucleus; (ii) while nuclear DP-rcDNA is sensitive to DNase I digestion, a small fraction of cytoplasmic DP-rcDNA is DNase I resistant; (iii) both DNase I-sensitive and -resistant cytoplasmic DP-rcDNAs cosediment with capsids and can be immunoprecipitated with HBV core antibody; and (iv) a primer extension assay maps the 5' end of the minus strand of DP-rcDNA at the authentic end of virion rcDNA. Hence, our results favor a hypothesis that the removal of viral polymerase protein covalently linked to the 5' end of the minus-strand DNA occurs inside the capsid in the cytoplasm and most possibly via a reaction that cleaves the phosphodiester bond between the tyrosine of the polymerase and the 5' phosphoryl group of minus-strand DNA.
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Details
- Title
- Characterization of the intracellular deproteinized relaxed circular DNA of hepatitis B virus: An intermediate of covalently closed circular DNA formation
- Creators
- Haitao Guo - Drexel UniversityDong Jiang - Drexel UniversityTianlun Zhou - Hepatitis B FoundationAndrea Cuconati - Hepatitis B FoundationTimothy M. Block - Drexel UniversityJu-Tao Guo - Drexel University
- Publication Details
- Journal of virology, v 81(22), pp 12472-12484
- Publisher
- Amer Soc Microbiology
- Number of pages
- 13
- Resource Type
- Journal article
- Language
- English
- Academic Unit
- Microbiology and Immunology
- Web of Science ID
- WOS:000254065400038
- Scopus ID
- 2-s2.0-36049013787
- Other Identifier
- 991019169552504721
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- Web of Science research areas
- Virology