title: Characterization of novel hepadnaviral RNA species accumulated in hepatoma cells treated with viral DNA polymerase inhibitors.
abstract: Inhibitors of hepadnaviral DNA polymerases are predicted to inhibit both minus and plus strand of viral DNA synthesis and arrest viral DNA replication at the stage of pregenomic (pg) RNA-containing nucleocapsids. However, analyses of the RNA species of human and duck hepatitis B viruses (HBV and DHBV, respectively) in hepatoma cells treated with viral DNA polymerase inhibitors revealed the genesis of novel RNA species migrating slightly faster than the full-length pgRNA. The DNA polymerase inhibitor-induced accumulation of these RNA species were abolished in the presence of alpha-interferon or HBV nucleocapsid assembly inhibitors. Moreover, they were protected from microccocal nuclease digestion and devoid of a poly-A tail. These characteristics suggest that the novel RNA species are most likely generated from RNase H cleavage of encapsidated pgRNA, after primer translocation and synthesis of the 5' terminal portion of minus strand DNA. In support of this hypothesis, DNA polymerase inhibitor treatment of chicken hepatoma cells transfected with a DHBV genome encoding an RNase H inactive DNA polymerase (E696H) failed to produce such RNA species. Our results thus suggest that the currently available DNA polymerase inhibitors do not efficiently arrest minus strand DNA synthesis at the early stage in hepatocytes. Hence, development of novel antiviral agents that more potently suppress viral DNA synthesis or viral nucleocapsid assembly inhibitors that are mechanistically complementary to the currently available DNA polymerase inhibitors are warranted.
authors.affiliation: Jiangsu Key Laboratory of New Drug Screening & Jiangsu Center for Pharmacodynamics Research and Evaluation & State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China; Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA.
authors.forename: Pinghu
authors.initials: P
authors.lastname: Zhang
authors.affiliation: Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA.
authors.forename: Fei
authors.initials: F
authors.lastname: Liu
authors.affiliation: Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA.
authors.forename: Fang
authors.initials: F
authors.lastname: Guo
authors.affiliation: Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA.
authors.forename: Qiong
authors.initials: Q
authors.lastname: Zhao
authors.affiliation: Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA.
authors.forename: Jinhong
authors.initials: J
authors.lastname: Chang
authors.affiliation: Baruch S. Blumberg Institute, Hepatitis B Foundation, Doylestown, PA, USA. Electronic address: ju-tao.guo@bblumberg.org.
authors.forename: Ju-Tao
authors.initials: JT
authors.lastname: Guo
mesh_terms: D000998:Antiviral Agents / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular; D002460:Cell Line; D004261:DNA Replication / Q000187:drug effects; D004279:DNA, Viral / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D015611:Hepatitis B Virus, Duck / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D006515:Hepatitis B virus / Q000187:drug effects / Q000235:genetics* / Q000502:physiology; D006801:Humans; D016898:Interferon-alpha / Q000494:pharmacology; D008113:Liver Neoplasms; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology*; D019251:Nucleocapsid / Q000378:metabolism; D012313:RNA / Q000378:metabolism; D012367:RNA, Viral / Q000302:isolation & purification / Q000378:metabolism*; D016914:Ribonuclease H / Q000378:metabolism; D019065:Virus Assembly / Q000187:drug effects; D014779:Virus Replication / Q000187:drug effects
chemical_list: D000998:Antiviral Agents; D004279:DNA, Viral; D016898:Interferon-alpha; D019384:Nucleic Acid Synthesis Inhibitors; D012367:RNA, Viral; C100808:pgRNA; D012313:RNA; D004259:DNA-Directed DNA Polymerase; D016914:Ribonuclease H