title: Enhanced pregenomic RNA levels and lowered precore mRNA transcription efficiency in a genotype A hepatitis B virus genome with C1766T and T1768A mutations obtained from a fulminant hepatitis patient.
abstract: The viral factors associated with the development of fulminant hepatitis B are not fully understood. We recently found four unique mutations [G to A at nucleotide 1742 (G1742A), C1766T, T1768A and T1809C] in the basal core promoter (BCP) region of a genotype A hepatitis B virus (HBV) strain (FH) obtained from a 53-year-old man with fatal fulminant hepatitis. To elucidate the association of the mutations of the FH genome with the disease, we constructed a 1.3-fold FH genome and its five variants by replacing one or two mutated nucleotides with wild-type nucleotide(s) via site-directed mutagenesis, and transfected human hepatoma cells (HepG2/C3A) with the constructs. There were no discernible differences between FH and two variants (FH_A1742G and FH_C1809T) with regard to viral replication and protein expression. However, in comparison to three other variants (FH_T1766C, FH_A1768T and FH_T1766C/A1768T) with wild-type nucleotide(s) at 1766 and/or 1768, the FH genome exhibited a 2.5-5-fold enhancement of viral replication by heightened pregenomic RNA synthesis and a 1.5-2.5-fold reduction in the hepatitis B e antigen (HBeAg) synthesis by the downregulation of the precore mRNA level. An immunofluorescence analysis revealed the increased and predominant cytoplasmic localization of the core protein in the FH genome. The present study demonstrates that the C1766T/T1768A mutations in the BCP region of genotype A HBV enhance viral replication, downregulate HBeAg expression and are responsible for the predominant localization of the core protein in the cytoplasm, which are likely associated with the development of fulminant hepatitis.
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi 329-0498, Japan.
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Nishizawa
authors.affiliation: Department of Gastroenterology, National Hospital Organization Takasaki General Medical Center, Gunma 370-0829, Japan.
authors.forename: Takashi
authors.initials: T
authors.lastname: Hoshino
authors.affiliation: Department of Gastroenterology, National Hospital Organization Takasaki General Medical Center, Gunma 370-0829, Japan.
authors.forename: Atsushi
authors.initials: A
authors.lastname: Naganuma
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi 329-0498, Japan.
authors.forename: Tominari
authors.initials: T
authors.lastname: Kobayashi
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi 329-0498, Japan.
authors.forename: Shigeo
authors.initials: S
authors.lastname: Nagashima
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi 329-0498, Japan.
authors.forename: Masaharu
authors.initials: M
authors.lastname: Takahashi
authors.affiliation: Department of Gastroenterology, National Hospital Organization Takasaki General Medical Center, Gunma 370-0829, Japan.|Department of Gastroenterology and Hepatology, Kusunoki Hospital, Gunma 375-0024, Japan.
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Takagi
authors.affiliation: Division of Virology, Department of Infection and Immunity, Jichi Medical University School of Medicine, Tochigi 329-0498, Japan.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Okamoto
mesh_terms: D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics*; D017809:Fatal Outcome; D016679:Genome, Viral; D023281:Genomics; D005838:Genotype; D006509:Hepatitis B / Q000821:virology*; D006515:Hepatitis B virus / Q000145:classification / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D008297:Male; D008875:Middle Aged; D008969:Molecular Sequence Data; D017354:Point Mutation; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000235:genetics*; D014779:Virus Replication
chemical_list: D004279:DNA, Viral; D012333:RNA, Messenger