title: RNA-templated replication of hepatitis delta virus: genomic and antigenomic RNAs associate with different nuclear bodies.
abstract: Lacking an RNA-dependent RNA polymerase, hepatitis delta virus (HDV), which contains a circular RNA of 1.7 kilobases, is nonetheless able to replicate its RNA by use of cellular transcription machineries. Previously, we have shown that the replications of genomic- and antigenomic-strand HDV RNAs have different sensitivities to alpha-amanitin, suggesting that these two strands are synthesized in different transcription machineries in the cells, but the nature of these transcription machineries is not clear. In this study, we performed metabolic labeling and immunofluorescence staining of newly synthesized HDV RNA with bromouridine after HDV RNA transfection into hepatocytes and confirmed that HDV RNA synthesis had both alpha-amanitin-sensitive and -resistant components. The antigenomic RNA labeling was alpha-amanitin resistant and localized to the nucleolus. The genomic RNA labeling was alpha-amanitin sensitive and more diffusely localized in the nucleoplasm. Most of the genomic RNA labeling appeared to colocalize with the PML nuclear bodies. Furthermore, promyelocytic leukemia protein, RNA polymerase II (Pol II), and the Pol I-associated transcription factor SL1 could be precipitated together with hepatitis delta antigen, suggesting the association of HDV replication complex with the Pol I and Pol II transcription machineries. This conclusion was further confirmed by an in vitro replication assay. These findings provide additional evidence that HDV RNA synthesis occurs in the Pol I and Pol II transcription machineries, thus extending the capability of the cellular DNA-dependent RNA polymerases to utilizing RNA as templates.
authors.affiliation: Department of Molecular Microbiology and Immunology, Keck School of Medicine, University of Southern California, 2011 Zonal Ave., Los Angeles, CA 90033-1054, USA.
authors.forename: Yi-Jia
authors.initials: YJ
authors.lastname: Li
authors.forename: Thomas
authors.initials: T
authors.lastname: Macnaughton
authors.forename: Lu
authors.initials: L
authors.lastname: Gao
authors.forename: Michael M C
authors.initials: MM
authors.lastname: Lai
mesh_terms: D000546:Amanitins / Q000494:pharmacology; D045744:Cell Line, Tumor; D002474:Cell-Free System / Q000378:metabolism; D016679:Genome, Viral / Q000502:physiology*; D006367:HeLa Cells; D003698:Hepatitis Delta Virus / Q000502:physiology*; D035923:Hepatitis delta Antigens / Q000096:biosynthesis; D022781:Hepatocytes / Q000378:metabolism / Q000821:virology; D006801:Humans; D047468:Immunoprecipitation; D042581:Intranuclear Space / Q000378:metabolism* / Q000821:virology; D008856:Microscopy, Fluorescence; D009363:Neoplasm Proteins / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism; D019384:Nucleic Acid Synthesis Inhibitors / Q000494:pharmacology; D035463:Pol1 Transcription Initiation Complex Proteins / Q000378:metabolism; D000071156:Promyelocytic Leukemia Protein; D012313:RNA / Q000096:biosynthesis* / Q000235:genetics; D012318:RNA Polymerase I / Q000378:metabolism; D012319:RNA Polymerase II / Q000037:antagonists & inhibitors / Q000378:metabolism; D016372:RNA, Antisense / Q000096:biosynthesis / Q000235:genetics; D000079962:RNA, Circular; D012367:RNA, Viral / Q000096:biosynthesis* / Q000235:genetics; D014157:Transcription Factors / Q000378:metabolism; D014158:Transcription, Genetic / Q000187:drug effects / Q000502:physiology; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D014779:Virus Replication / Q000187:drug effects / Q000502:physiology*
chemical_list: D000546:Amanitins; D035923:Hepatitis delta Antigens; D009363:Neoplasm Proteins; D009687:Nuclear Proteins; D019384:Nucleic Acid Synthesis Inhibitors; D035463:Pol1 Transcription Initiation Complex Proteins; D000071156:Promyelocytic Leukemia Protein; D016372:RNA, Antisense; D000079962:RNA, Circular; D012367:RNA, Viral; C091129:TAF1B protein, human; D014157:Transcription Factors; D025521:Tumor Suppressor Proteins; C070576:PML protein, human; D012313:RNA; D012319:RNA Polymerase II; D012318:RNA Polymerase I