1887

Abstract

We recently established a cell culture system for the replication of hepatitis C virus (HCV) by using a human T cell leukaemia virus type I-infected cell line, MT-2, and showed that the quasi-species of the hypervariable region 1 observed in the original inoculum became homogeneous in MT-2 cells 10 days after inoculation of HCV. In this study, we obtained HCV cDNA clones covering the whole viral genome by RT-nested PCR using RNA from HCV- infected cloned MT-2C cells, which support viral replication more efficiently, at 12 days after inoculation. A total of 41 distinct HCV cDNA clones covering almost the whole viral genome were also isolated from a cDNA library derived from the original inoculum. Molecular evolutional analyses comparing the sequences of the HCV clones obtained from both sources revealed that the HCV populations became homogeneous in more than half of the compared regions. This finding suggests that limited HCV populations are able to replicate in MT-2C cells. In addition, we isolated cDNA clones containing a 3′ X-tail sequence, which was recently identified as a bona fide 3′ terminus of the HCV genome, in the HCV-infected MT-2C cells and confirmed that the nucleotide sequence of the 3′ X-tail was highly conserved, suggesting its implication in HCV replication. Finally, on the basis of the sequences of HCV cDNA clones obtained from HCV- infected MT-2C cells, we determined the entire nucleotide sequence of the HCV genome containing the 3′ X-tail as a candidate for an infectious HCV molecular clone.

Loading

Article metrics loading...

/content/journal/jgv/10.1099/0022-1317-78-2-329
1997-02-01
2024-12-12
Loading full text...

Full text loading...

/deliver/fulltext/jgv/78/2/9018054.html?itemId=/content/journal/jgv/10.1099/0022-1317-78-2-329&mimeType=html&fmt=ahah

References

  1. Bukh J., Miller R. H., Purcell R. H. 1995; Genetic heterogeneity of hepatitis C virus: quasispecies and genotypes. [Review]. Seminars in Liver Disease 15:41–63
    [Google Scholar]
  2. Chen P. J., Lin M. H., Tai K. F., Liu P. C., Lin C. J., Chen D. S. 1992; The Taiwanese hepatitis C virus genome : sequence determination and mapping the 5′ termini of viral genomic and antigenomic RNA. Virology 188:102–113
    [Google Scholar]
  3. Cribier B., Schmitt C., Bingen A., Kirn A., Keller F. 1995; In vitro infection of peripheral blood mononuclear cells by hepatitis C virus. Journal of General Virology 76:2485–2491
    [Google Scholar]
  4. Hijikata M., Mizuno K., Rikihisa T., Shimizu Y. K., Iwamoto A., Nakajima N., Yoshikura H. 1995; Selective transmission of hepatitis C virus in vivo and in vitro. Archives of Virology 140:1623–1628
    [Google Scholar]
  5. Honda M., Kaneko S., Unoura M., Kobayashi K., Murakami S. 1992; Sequence comparisons for a hepatitis C virus genome RNA isolated from a patient with liver cirrhosis. Gene 120:317–318
    [Google Scholar]
  6. Kato N., Hijikata M., Ootsuyama Y., Nakagawa M., Ohkoshi S., Shimotohno K. 1990a; Sequence diversity of hepatitis C viral genomes. Molecular Biology & Medicine 7:495–501
    [Google Scholar]
  7. Kato N., Hijikata M., Ootsuyama Y., Nakagawa M., Ohkoshi S., Sugimura T., Shimotohno K. 1990b; Molecular cloning of the human hepatitis C virus genome from Japanese patients with non-A, non- B hepatitis. Proceedings of the National Academy of Sciences USA: 879524–9528
    [Google Scholar]
  8. Kato N., Hijikata M., Nakagawa M., Ootsuyama Y., Muraiso K., Ohkoshi S., Shimotohno K. 1991; Molecular structure of the Japanese hepatitis C viral genome. FEBS Letters 280:325–328
    [Google Scholar]
  9. Kato N., Ootsuyama Y., Ohkoshi S., Nakazawa T., Sekiya H., Hijikata M., Shimotohno K. 1992a; Characterization of hypervariable regions in the putative envelope protein of hepatitis C virus. Biochemical and Biophysical Research Communications 189:119–127
    [Google Scholar]
  10. Kato N., Ootsuyama Y., Tanaka T., Nakagawa M., Nakazawa T., Muraiso K., Ohkoshi S., Hijikata M., Shimotohno K. 1992b; Marked sequence diversity in the putative envelope proteins of hepatitis C viruses. Virus Research 22:107–123
    [Google Scholar]
  11. Kato N., Sekiya H., Ootsuyama Y., Nakazawa T., Hijikata M., Ohkoshi S., Shimotohno K. 1993; Humoral immune response to hypervariable region 1 of the putative envelope glycoprotein (gp70) of hepatitis C virus. Journal of Virology 67:3923–3930
    [Google Scholar]
  12. Kato N., Ootsuyama Y., Sekiya H., Ohkoshi S., Nakazawa T., Hijikata M., Shimotohno K. 1994; Genetic drift in hypervariable region 1 of the viral genome in persistent hepatitis C virus infection. Journal of Virology 68:4776–4784
    [Google Scholar]
  13. Kato N., Nakazawa T., Mizutani T., Shimotohno K. 1995; Susceptibility of human T-lymphotropic virus type I infected cell line MT-2 to hepatitis C virus infection. Biochemical and Biophysical Research Communications 206:863–869
    [Google Scholar]
  14. Kato N., Ikeda M., Mizutani T., Sugiyama K., Noguchi M., Hirohashi S., Shimotohno K. 1996; Replication of hepatitis C virus in cultured non-neoplastic human hepatocytes. Japanese Journal ofCancer Research 87:787–792
    [Google Scholar]
  15. Khromykh A. A., Westaway E. G. 1994; Completion of Kunjin virus RNA sequence and recovery of an infectious RNA transcribed from stably cloned full-length cDNA. Journal of Virology 68:4580–4588
    [Google Scholar]
  16. Lai C. J., Zhao B. T., Hori H., Bray M. 1991; Infectious RNA transcribed from stably cloned full-length cDNA of dengue type 4 virus. Proceedings of the National Academy of Sciences USA: 885139–5143
    [Google Scholar]
  17. Lanford R. E., Sureau C., Jacob J. R., White R., Fuerst T. R. 1994; .Demonstration of in vitro infection of chimpanzee hepatocytes with hepatitis C virus using strand-specific RT/PCR. Virology 202:606–614
    [Google Scholar]
  18. Miller R. H., Purcell R. H. 1990; Hepatitis C virus shares amino acid sequence similarity with pestiviruses and flaviviruses as well as members of two plant virus supergroups. Proceedings of the National Academy of Sciences USA: 872057–2061
    [Google Scholar]
  19. Mizutani T., Kato N., Hirota M., Sugiyama K., Murakami A., Shimotohno K. 1995; Inhibition of hepatitis C virus replication by antisense oligonucleotide in culture cells. Biochemical and Biophysical Research Communications 212:906–911
    [Google Scholar]
  20. Mizutani T., Kato N., Saito S., Ikeda M., Sugiyama K., Shimotohno K. 1996; Characterization of hepatitis C virus replication in cloned cells obtained from a human T-cell leukemia virus type I infected cell line, MT- 2. Journal of Virology 70:7219–7223
    [Google Scholar]
  21. Moormann R. J. M., Genip H. G. P., Miedema G. K. W., Hulst M. M., Rijn P. A. 1996; Infectious RNA transcribed from an engineered full- length cDNA template of the genome of a pestivirus. Journal of Virology 70:763–770
    [Google Scholar]
  22. Nakajima N., Hijikata M., Yoshikura H., Shimizu Y. K. 1996; .Characterization of long-term cultures of hepatitis C virus. Journal of Virology 70:3325–3329
    [Google Scholar]
  23. Nei M., Gojobori T. 1986; Simple methods for estimating the numbers of synonymous and nonsynonymous nucleotide substitutions. Molecular Biology and Evolution 3:418–426
    [Google Scholar]
  24. Noguchi M., Hirohashi S. 1996; Cell lines from non-neoplastic liver and hepatocellular carcinoma tissue from a single patient. In Vitro Cellular & Developmental Biology Animal 32:135–137
    [Google Scholar]
  25. Ohkoshi S., Kojima H., Tawaraya H., Miyajima T., Kamimura T., Asakura H., Satoh A., Hirose S., Hijikata M., Kato N., Shimotohno K. 1990; Prevalence of antibody against non-A, non-B hepatitis virus in Japanese patients with hepatocellular carcinoma. Japanese Journal of Cancer Research 81:550–553
    [Google Scholar]
  26. Rice C. M., Grakoui A., Galler R., Chambers T. J. 1989; Transcription of infectious yellow fever RNA from full-length cDNA templates produced by in vitro ligation. New Biologist 1:285–296
    [Google Scholar]
  27. Saito I., Miyamura T., Ohbayashi A., Harada H., Katayama T., Kikuchi S., Watanabe Y., Koi S., Onji M., Ohta Y., Choo Q. -L., Houghton M., Kuo G. 1990; Hepatitis C virus infection is associated with the development of hepatocellular carcinoma. Proceedings of the National Academy of Sciences USA: 87 6547–6549
    [Google Scholar]
  28. Saito S., Kato N., Hijikata M., Gunji T., Itabashi M., Kondo M., Tanaka K., Shimotohno K. 1996; Comparison of hypervariable regions (HVR1 and HVR2) in positive- and negative-stranded hepatitis C virus RNA in cancerous and non-cancerous liver tissue, peripheral blood mononuclear cells and serum from a patient with hepatocellular carcinoma. International Journal of Cancer 67: 199–203
    [Google Scholar]
  29. Sekiya H., Kato N., Ootsuyama Y., Nakazawa T., Yamauchi K., Shimotohno K. 1994; Genetic alterations of the putative envelope proteins encoding region of the hepatitis C virus in the progression to relapsed phase from acute hepatitis: humoral immune response to hypervariable region 1. International Journal of Cancer 57: 664–670
    [Google Scholar]
  30. Shimizu Y. K., Purcell R. H., Yoshikura H. 1993; Correlation between the infectivity of hepatitis C virus in vivo and its infectivity in vitro. Proceedings of the National Academy of Sciences USA: 906037–6041
    [Google Scholar]
  31. Shimotohno K. 1995; Hepatitis C virus as a causative agent of hepatocellular carcinoma. Intervirology 38:162–169
    [Google Scholar]
  32. Simmonds P. 1995; Variability of hepatitis C virus. Review Hepatology 21:570–583
    [Google Scholar]
  33. Sumiyoshi H., Hoke C. H., Trent D. W. 1992; Infectious Japanese encephalitis virus RNA can be synthesized from in vitro-ligated cDNA templates. Journal of Virology 66:5425–5431
    [Google Scholar]
  34. Takamizawa A., Mori C., Fuke I., Manabe S., Murakami S., Fujita J., Onishi E., Andoh T., Yoshida I., Okayama H. 1991; Structure and organization of the hepatitis C virus genome isolated from human carriers. Journal of Virology 65:1105–1113
    [Google Scholar]
  35. Tanaka T., Kato N., Nakagawa M., Ootsuyama Y., Cho M. J., Nakazawa T., Hijikata M., Ishimura Y., Shimotohno K. 1992; Molecular cloning of hepatitis C virus genome from a single Japanese carrier: sequence variation within the same individual and among infected individuals. Virus Research 23:39–53
    [Google Scholar]
  36. Tanaka T., Kato N., Cho M. J., Shimotohno K. 1995; A novel sequence found at the 3′ terminus ofhepatitis C virus genome. Biochemical and Biophysical Research Communications 215:744–749
    [Google Scholar]
  37. Tanaka T., Kato N., Cho M. J., Sugiyama K., Shimotohno K. 1996; Structure of the 3′ terminus of the hepatitis C virus genome. Journal of Virology 70:3307–3312
    [Google Scholar]
  38. Weiner A. J., Geysen H. M., Christopherson C., Hall J. E., Mason T. J., Saracco G., Bonino F., Crawford K., Marion C. D., Crawford K. A., Brunetto M., Bar P. J., Miyamuta T., McHutchinson J., Houghton M. 1992; Evidence for immune selection of hepatitis C virus (HCV) putative envelope glycoprotein variants : potential role in chronic HCV infections. Proceedings of the National Academy of Sciences USA: 893468–3472
    [Google Scholar]
  39. Zibert A., Schreier E., Roggendorf M. 1995; Antibodies in human sera specific to hypervariable region 1 of hepatitis C virus can block viral attachment. Virology 208:653–661
    [Google Scholar]
/content/journal/jgv/10.1099/0022-1317-78-2-329
Loading
/content/journal/jgv/10.1099/0022-1317-78-2-329
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error