Molecular cloning of a defective hepatitis C virus genome from the ascitic fluid of a patient with hepatocellular carcinoma.
A defective hepatitis C virus (HCV) genome in the ascitic fluid of a patient with hepatocellular carcinoma was cloned and sequenced up to the 3' poly(U) stretch. When compared with the published Taiwanese HCV sequence, this defective genome contained deletions of single nucleotides at eight sites, double nucleotides at two sites, triple nucleotides at four sites, quadruple nucleotides at one site and replacement of a short stretch of sequence at one site. For comparison, the corresponding regions containing these mutations were also cloned from a serum sample from this patient. Except for deletions of two triple nucleotides in the hypervariable region, the reading frames of all serum-derived clones were intact. The defective HCV genome encoded a truncated core protein with 90 amino acid residues (the last 20 amino acid residues came from a different reading frame), whereas the serum-derived genome encoded a full-length core protein. When expressed in Huh-7 cells, these two proteins were localized to the nucleus and cytoplasm, respectively. Using specific primer-sets, ascites- and serum-derived genomes were each detected alone in ascitic fluid and serum samples, respectively, whereas both sequences were present in ascitic mononuclear cells. The defective sequence thus constituted the major virus population in the ascitic fluid whereas a putative helper genome coexisted with it inside the ascitic mononuclear cells. This sequence is possibly a defective and interfering genome.
Yeh
C T
CT
Liver Research Unit, Chang-Gung Memorial Hospital and Medical College, Taipei, Taiwan.
Lu
S C
SC
Chu
C M
CM
Liaw
Y F
YF
D000595:Amino Acid Sequence; D001202:Ascitic Fluid / Q000821:virology*; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000821:virology*; D003001:Cloning, Molecular; D016679:Genome, Viral; D006801:Humans; D008113:Liver Neoplasms / Q000235:genetics / Q000821:virology*; D008969:Molecular Sequence Data; D016415:Sequence Alignment; D017421:Sequence Analysis