Structure and Function of the Hepatitis C Virus Internal Ribosome Entry Site

  • C. Wang
  • A. Siddiqui
Part of the Current Topics in Microbiology and Immunology book series (CT MICROBIOLOGY, volume 203)


Human hepatitis C virus (HCV) infects hepatocytes and the viral infection often develops into chronic disease, liver cirrhosis and hepatocellular carcinoma (Saito et al. 1990; Plagemann 1991; Ruiz et al. 1992). Currently HCV infections account for about 80% of posttransfusion hepatitis worldwide. To date HCV has not been successfully grown in any cultured cells (Shimizu et al. 1993) which precludes the biological studies of this virus. The discovery of the human HCV stems from an unprecedented use of molecular cloning techniques rather than by the conventional method of virus isolation (Choo et al. 1989). Nucleic acids extracted from the sera of chimpanzees infected with infectious non-A, non-B hepatitis plasma were usedto prepare a λ gt11 library which was then immunoscreened with non-A, non-B hepatitis serum (Choo et al. 1989). The viral RNA genome cloned as a cDNA was characterized by nucleotide sequence determination to uncover the genetic order of the coding potential. Further comparison of the complete HCV genome to other known viral genomes placed this virus in the family of Flaviviridae (Miller and Purcell 1990; Houghton et al. 1991). Members of this virus group include human flaviviruses (e.g., Yellow fever virus, Dengue virus and St. Louis encephalitis virus) and animal pestiviruses (e.g., bovine viral diarrhea virus and hog cholera virus).


Internal Ribosome Entry Site Bovine Viral Diarrhea Virus InfectiOUs Bronchitis VIrUS Chloramphenicol Acetyl Transferase Internal Ribosome Entry Site Element 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


  1. Agol VI (1991) The 5’-untranslated region of picornaviral genomes. Adv Virus Res 40: 103–180PubMedCrossRefGoogle Scholar
  2. Brierley IN, Digard P, Inglis SC (1989) Characterization of an efficient coronavirus ribosomal frame- shifting signal: requirement for an RNA pseudoknot. Cell 57: 537–547PubMedCrossRefGoogle Scholar
  3. Brierley IN, Rolley J, Jenner JA, Inglis SC (1991) Mutational analysis of the RNA pseudoknot component of a coronavirus ribosomal frameshifting signal. J Mol Biol 220: 889–902PubMedCrossRefGoogle Scholar
  4. Brock KV, Deng R, Riblet SM (1992) Nucleotide sequencing of 5’ and 3’ termini of bovine viral diarrhea virus by RNA ligation and PCR. J Virol Methods 38: 39–46PubMedCrossRefGoogle Scholar
  5. Brown EA, Day SP, Jansen RW, Lemon SM (1991) The 5’ nontranslated region of hepatitis A virus RNA: secondary structure and elements required for translation in vitro. J Virol 65; 5825–5838Google Scholar
  6. Brown EA, Zhang H, Ping L-H, Lemon SM (1992) Secondary structure of the 5’ nontranslated regions of hepatitis C virus and pestivirus genomic RNAs. Nucleic Acids Res 20: 5041–5045PubMedCrossRefGoogle Scholar
  7. Brown EA, Zajac AJ, Lemon SM (1994) In vitro characterization of an internal ribosomal entry site (IRES) present within the 5’ nontranslated region of hepatitis A virus: comparison with the IRES of encephalomyocarditis virus. J Virol 68: 1066–1074PubMedGoogle Scholar
  8. Bukh J, Purcell RH, Miller RH (1992) Sequence analysis of the 5’ noncoding region of hepatitis C virus. Proc Natl Acad Sci USA 89: 4942–4946PubMedCrossRefGoogle Scholar
  9. Chambers TJ, Hahn CS, Galler R, Rice CM (1990) Flavivirus genome organization, expression, and replication. Annu Rev Microbiol 44: 649–688PubMedCrossRefGoogle Scholar
  10. Chamorro M, Parkin N, Varmus HE (1992) An RNA pseudoknot and an optimal heptameric shift site are required for high efficient ribosomal frameshifting on a retroviral messenger RNA. Proc Natl Acad Sci USA 89: 713–717PubMedCrossRefGoogle Scholar
  11. Chen P-J, Lin M-H, Tai K-F, Liu P-C, Li C-J, Chen K-S (1992) The Taiwanese hepatitis C virus genome: sequence determination and mapping the 5’ termini of viral genomic and antigenomic RNA. Virology 188: 102–113PubMedCrossRefGoogle Scholar
  12. Choo Q-L, Kuo G, Weiner AJ, Overby LR, Bradley DW, Houghton M (1989) Isolation of a cDNA fragment from a blood-borne non-A, non-B viral hepatitis agent. Science 244: 359–362PubMedCrossRefGoogle Scholar
  13. Choo Q-L, Richman KH, Han JH, Berger K, Lee C, Dong C, Gallegos C, Coit D, Medina-Selby A, Barr PJ, Weiner AJ, Bradley DW, Kuo G, Houghton M (1991) Genetic organization and diversity of the hepatitis C virus. Proc Natl Acad Sci USA 88: 2451–2455PubMedCrossRefGoogle Scholar
  14. Collett MS, Larson R, Gold C, Strick D, Anderson DK, Purchio AF (1988) Molecular cloning and nucleotide sequence of the pestivirus bovine viral diarrhea virus. Virology 165: 191–199CrossRefGoogle Scholar
  15. Collett MS, Anderson DK, Retzel E (1989a) Comparison of the pestivirus bovine viral diarrhea virus with members of flaviviridae. J Gen Virol 69: 2637–2643CrossRefGoogle Scholar
  16. Collett MS, Moenning V, Horzinek M (1989b) Recent advances in pestivirus research. J Gen Virol 70: 253–266PubMedCrossRefGoogle Scholar
  17. Deng R, Brock KV (1993) 5’ and 3’untranslated regions of pestivirus genome: primary and secondary structure analyses. Nucleic Acids Res 21: 1949–1957PubMedCrossRefGoogle Scholar
  18. Duke GM, Hoffman MA, Palmenberg AC (1992) Sequence and structural elements that contribute to efficient encephalomyocarditis virus RNA translation. J Virol 66: 1602–1609PubMedGoogle Scholar
  19. Feng YX, Yuan H, Rein A, Levin JG (1992) Bipartite signal for read-through suppression in murine leukemia virus mRNA: an eight-nucleotide purine-rich sequence immediately downstream of the gag termination codon followed by an RNA pseudoknot. J Virol 66: 5127–5132PubMedGoogle Scholar
  20. Glass MJ, Summers DF (1993) Identification of a trans-acting activity from liver that stimulates hepatitis A virus translation in vitro. Virology 193: 1047–1050PubMedCrossRefGoogle Scholar
  21. Glass MJ, Jia X-Y, Summers DF (1993) Identification of the hepatitis A virus internal ribosome entry site: in vivo and in vitro analysis of bicistronic RNAs containing the HAV 5’ noncoding region. Virology 193: 842–852PubMedCrossRefGoogle Scholar
  22. Grakoui A, Wychowski C, Lin C, Feinstone SM, Rice CM (1993) Expression and identification of hepatitis C virus polyprotein cleavage products. J Virol 67: 1385–1395PubMedGoogle Scholar
  23. Han JH, Houghton M (1992) Group specific sequences and conserved secondary structures at the 3’ end of HCV genome and its implication for viral replication. Nucleic Acids Res 20: 3520PubMedCrossRefGoogle Scholar
  24. Han JH, Shyamala V, Richman KH, Brauer MJ, Irvine B, Urdea MS, Tekamp-Olson P, Kuo G, Choo Q-L, Houghton M (1991) Characterization of the terminal regions of hepatitis C viral RNA: identification of conserved sequences in the 5’ untranslated region and poly(A) tails at the 3’ end. Proc Natl Acad USA 88: 1711–1715CrossRefGoogle Scholar
  25. Hijikata M, Mizushima H, Tanji Y, Komoda Y, Hirowatari T, Akagi T, Kato N, Kimura K, Shimotohno K (1993) Proteolytic processing and membrane association of putative nonstructural proteins of hepatitis C virus. Proc Natl Acad Sci USA 90: 10773–10777PubMedCrossRefGoogle Scholar
  26. Houghton M, Weiner A, Han J, Kuo G, Choo Q-L (1991) Molecular biology of the hepatitis C viruses: implications for diagnosis, development and control of viral disease. Hepatology 14: 381–388PubMedCrossRefGoogle Scholar
  27. Iizuka N, Najita L, Franzusoff A, Sarnow P (1994) Cap-dependent and cap-independent translation of mRNA in cell-free extracts prepared from Saccharomyces cerevisiae. Mol Cell Biol (in press)Google Scholar
  28. Inchauspe G, Zebedee S, Lee D-H, Sugitani M, Nasoff M, Prince AM (1991) Genomic structure of the human prototype strain H of hepatitis C virus: comparison with American and Japanese isolates. Proc Natl Acad Sci USA 88: 10292–10296PubMedCrossRefGoogle Scholar
  29. Jackson RJ (1991) Potassium salts influence the fidelity of mRNA translation initiation in rabbit reticulocyte lysates: unique features of encephalomyocarditis virus RNA translation. Biochim Biophys Acta 1088: 345–358PubMedGoogle Scholar
  30. Jackson RJ, Howell MT, Kaminiski A (1990) The novel mechanism of initiation of picornavirus RNA translation. Trends Biochem Sci 15: 477–483PubMedCrossRefGoogle Scholar
  31. Jang SK, Wimmer E (1990) Cap-independent translation of encephalomyocarditis virus RNA: structural elements of the internal ribosomal entry site and involvement of a cellular 57-KD RNA-binding protein. Genes Dev 4: 1560–1572PubMedCrossRefGoogle Scholar
  32. Jang SK, Davies MV, Kaufman RJ, Wimmer E (1989) Initiation of protein synthesis by internal entry of ribosomes into the 5’ nontranslated region of encephalomyocarditis virus RNA in vivo. J Virol 63: 1651–1660PubMedGoogle Scholar
  33. Jia X-Y, Scheper G, Brown D, Updike W, Harmon S, Richards D, Summers D, Erenfeld E (1991) Translation of hepatitis A virus RNA in vitro: aberrant internal initiations influenced by 5’ noncoding region. Virology 182: 712–722PubMedCrossRefGoogle Scholar
  34. Kaminski A, Howell MT, Jackson RJ (1990) Initiation of encephalomyocarditis virus RNA translation: the authentic initiation site is not selected by a scanning mechanism. EMBO J 9: 3753–3759PubMedGoogle Scholar
  35. Kaminski A, Belsham GJ, Jackson RJ (1994) Translation of encephalomyocarditis virus RNA: parameters influencing the selection of the internal initiation site. EMBO J 13: 1673–1681PubMedGoogle Scholar
  36. Kato N, Hijikata M, Ootsuyama Y, Nakagawa M, Ohkoshi S, Sugimura T, Shimotohno K (1990) Molecular cloning of the human hepatitis C virus genome from Japanese patients with non-A, non-B hepatitis. Proc Natl Acad Sci USA 87: 9524–9528PubMedCrossRefGoogle Scholar
  37. Kettinen H, Grace K, Grunert S, Clarke B, Rowlands D, Jackson RJ (1993) Mapping of the internal ribosome entry site at the 5’ end of the hepatitis C virus genome. Proceedings of the International Symposium on Viral Hepatitis and Liver Disease. Tokyo (In press)Google Scholar
  38. Koonin EV (1993) Computer-assisted identification of a putative methyltransferase domain in NS5 protein of flaviviruses and 12 protein of reovirus. J Gen Virol 74: 733–740PubMedCrossRefGoogle Scholar
  39. Kozak M (1992) Regulation of translation in eukaryotic system. Annu Rev Cell Biol 8: 197–225PubMedCrossRefGoogle Scholar
  40. Kuhn R, Luz N, Beck E (1990) Functional analysis of the internal translation initiation site of foot-and- mouth disease virus. J Virol 64: 4625–4631PubMedGoogle Scholar
  41. Le S-Y, Chen J-H, Sonenberg N, Maizel JV (1992) Conserved tertiary structure elements in the 5‘untranslated region of human enteroviruses and rhinoviruses. Virology 191: 858–866PubMedCrossRefGoogle Scholar
  42. Le S-Y, Chen J-H, Sonenberg N, Maizel JV Jr (1993) Conserved tertiary structural elements in the 5’ nontranslated region of cardiovirus, aphthovirus and hepatitis A virus RNAs. Nucleic Acids Res 21: 2445–2451PubMedCrossRefGoogle Scholar
  43. Le S-Y, Sonenberg N, Maizel JV Jr (1994) Distinct structural elements and internal entry of ribosomes in mRNA3 encoded by infectious bronchitis virus. Virology 198: 405–411PubMedCrossRefGoogle Scholar
  44. Lee KAW, Sonenberg N (1982) Inactivation of cap-binding proteins accompanies the shut-off of host protein synthesis by poliovirus. Proc Natl Acad Sci USA 79: 3447–3451PubMedCrossRefGoogle Scholar
  45. Leathers V, Tanguay R, Kobayashi M, Gallie DR (1993) A phylogenetically conserved sequence within viral 3’ untranslated RNA pseudoknots regulates translation. Mol Cell Biol 13: 5331–5347PubMedGoogle Scholar
  46. Liu DX, Inglis SC (1992) Internal entry of ribosomes on a tricistronic mRNA encoded by infectious bronchitis virus. J virol 66: 6143–6154PubMedGoogle Scholar
  47. Meerovitch K, Nicholson R, Sonenberg N (1991) In vitro mutational analysis of cis-acting RNA translational elements within the poliovirus type 2 5’ untranslated region. J Virol 65: 5895–5901PubMedGoogle Scholar
  48. Miller RH, Purcell RH (1990) Hepatitis C virus shares amino acid sequence similarity with pestiviruses and flaviviruses as well as members of two plant virus supergroups. Proc Natl Acad Sci USA 87: 2057–2061PubMedCrossRefGoogle Scholar
  49. Nicholson R, Pelletier J, Le S-Y, Sonenberg N (1991) Structural and functional analysis of the ribosome landing pad of poliovirus type 2: in vivo translational studies. J Virol 65: 5886–5894PubMedGoogle Scholar
  50. Okamoto H, Okada S, Sugiyama Y, Kurai K, Lizuka H, Machida A, Miyakawa Y, Mayumi M (1991) Nucleotide sequence of the genomic RNA of hepatitis C virus isolated from a human carrier: comparison with reported isolates for conserved and divergent regions. J Gen Virol 72: 2697–2704PubMedCrossRefGoogle Scholar
  51. Pelletier J, Sonenberg N (1988) Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature 334: 320–325PubMedCrossRefGoogle Scholar
  52. Percy N, Belsham GJ, Brangwyn JK, Sullivan M, Stone DM, Almond JW (1992) Intracellular modifications induced by poliovirus reduce the requirement for structural motifs in the 5’ noncoding region of the genome involved in internal initiation of protein synthesis. J Virol 66: 1695–1701PubMedGoogle Scholar
  53. Pestova TV, Hellen CUT, Wimmer E (1991) Translation of poliovirus RNA: role of an essential cis-acting oligopyrimidine element within the 5’ nontranslated region and involvement of a cellular 57-kilodalton protein. J Virol 65: 6194–6204PubMedGoogle Scholar
  54. Pilipenko EV, Blinov VM, Ramanova LI, Sinyakov AN, Maslova SV, Agol VI (1989) Conserved structural domains in the 5’-untranslated region of picornaviral genomes: an analysis of the segment controlling translation and neurovirulence. Virology 168: 201–209PubMedCrossRefGoogle Scholar
  55. Pilipenko EV, Gmyl AP, Maslova SV, Svitkin YV, Sinyakov AN, Agol VI (1992) Prokaryoticlike cis elements in the cap-independent internal initiation of translation on picornavirus RNA. Cell 68: 119–131PubMedCrossRefGoogle Scholar
  56. Plagemann PGW (1991) Hepatitis C virus. Arch Virol 120: 165–180PubMedCrossRefGoogle Scholar
  57. Pletnev AG, Yamshchikov VF, Blinov VM (1990) Nucleotide sequence of the genome and complete amino acid sequence of the polyprotein of tick-borne encephalitis virus. Virology 174: 250–263PubMedCrossRefGoogle Scholar
  58. Poole TL, Wang C, Popp RA, Potgieter LND, Siddiqui A, Collett MS (1995) Pestivirus translation initiation by internal ribosome entry. Virology 206: 750–754PubMedCrossRefGoogle Scholar
  59. Reynolds JE, Grace K, Clarke BE, Rowlands DJ, Kaminski A, Jackson R (1995) Unusual features of internal initiation of translation of hepatitis C virus RNA (submitted)Google Scholar
  60. Rice CM, Strauss EG, Strauss HJ (1986) Structure of the flavivirus genome. In: Schlessinger S, Schlessinger MJ (eds) The togaviridae and flaviviridae. Plenum, New York, pp 279–326Google Scholar
  61. Ruiz J, Sangro B, Cuende J I, Beloqui O, Riezu-Boj Jl, Herrero J I, Prieto J (1992) Hepatitis B and C viral infections in patients with hepatocellular carcinoma. Hepatology 16: 637–641PubMedCrossRefGoogle Scholar
  62. Saito I, Miyamura T, Ohbayashi A, Harada H, Katayama T, Kikuchi S, Watanabe S, Koi TY, Onji M, Ohta Y, Choo Q-L, Houghton M, Kuo G (1990) Hepatitis C virus infection is associated with the development of hepatocellular carcinoma. Proc Natl Acad Sci USA 87: 6547–6549PubMedCrossRefGoogle Scholar
  63. Schimmel P (1989) RNA pseudoknots that interact with components of the translation apparatus. Cell 58: 9–12PubMedCrossRefGoogle Scholar
  64. Selby MJ, Choo Q-L, Berger K, Kuo G, Glazer E, Eckart M, Lee C, Chien D, Kuo C, Houghton M (1993) Expression, identification and subcellular localization of the proteins encoded by the hepatitis C viral genome. J Gen Virol 74: 1103–1113PubMedCrossRefGoogle Scholar
  65. Shamoo Y, Tarn A, Konigsberg WH, Williams KR (1993) Translational repression by the bacteriophage T4 gene 32 protein involves specific recognition of an RNA pseudoknot structure. J Mol Biol 232: 89–104PubMedCrossRefGoogle Scholar
  66. Shih DS, Park l-W, Evans CL, Jaynes JM, Palmenberg AC (1987) Effects of cDNA hybridization on translation of encephalomyocarditis virus RNA. J Virol 61: 2033–2037PubMedGoogle Scholar
  67. Shimizu YK, Purcell RH, Yoshikura H (1993) Correlation between the infectivity of hepatitis C virus in vivo and its infectivity in vitro. Proc Natl Acad Sci USA 90: 6037–6041PubMedCrossRefGoogle Scholar
  68. Skinner MA, Racaniello VR, Dunn G, Cooper J, Minor PD, Almond JW (1989) New model for the secondary structure of the 5’ non-coding RNA of poliovirus is supported by biochemical and genetic data that also show that RNA secondary structure is important in neurovirulence. J Mol Biol 207: 379–392PubMedCrossRefGoogle Scholar
  69. Somogyi P, Jenner AJ, Brierley I, Inglis SC (1993) Ribosomal pausing during translation of an RNA pseudoknot. Mol Cell Biol 13: 6931–6940PubMedGoogle Scholar
  70. 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. J Virol 65: 1105–1113PubMedGoogle Scholar
  71. Tanaka T, Kato N, Nakagawa M, Ootruyama Y, Cho MJ, 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 Res 23: 39–53PubMedCrossRefGoogle Scholar
  72. Tsukiyama-Kohara K, lizuka N, Kohara M, Nomoto A (1992) Internal ribosome entry site within hepatitis C virus RNA. J Virol 66: 1476–1483PubMedGoogle Scholar
  73. Tu C, Tzeng T-H, Bruenn JH (1992) Ribosomal movement impeded at a pseudoknot required for frameshifting. Proc Natl Acad Sci USA 89: 8636–8640PubMedCrossRefGoogle Scholar
  74. Tzeng T-H, Tu C-L, Bruenn JA (1992) Ribosomal frameshifting require a pseudoknot in the Saccha- romyces cerevisiae double-stranded RNA virus. J Virol 66: 999–1006PubMedGoogle Scholar
  75. Wang C, Le SY, Siddiqui A (1995) Functional role of an RNA pseudoknot structure in internal initiation of translation of hepatitis C virus RNA genome, (submitted for publication)Google Scholar
  76. Wang C, Sarnow P, Siddiqui A (1993) Translation of human hepatitis C virus RNA in cultured cells in mediated by an internal ribosome-binding mechanism. J Virol 67: 3338–3344PubMedGoogle Scholar
  77. Wang C, Sarnow P, Siddiqui A (1994) A conserved helical element is essential for the internal initiation of translation of the hepatitis C virus RNA. J Virol 68: 7301–7307PubMedGoogle Scholar
  78. Wills NM, Gesteland RF, Atkins JF (1991) Evidence that a downstream pseudoknot is required for translational read-through of the moloney murine leukemia virus gag stop codon. Proc Natl Acad Sci USA 88: 6991–6995PubMedCrossRefGoogle Scholar
  79. Wimmer E, Hellen CUT, Cao X (1993) Genetics of poliovirus. Annu Rev Genet 27: 353–436PubMedCrossRefGoogle Scholar
  80. Wiskerchen M, Belzer SK, Collett MS (1991) Pestivirus gene expression: the first protein product of the bovine viral diarrhea virus large open reading frame, p20, possesses proteolytic activity. J Virol 65’ 4509–4514Google Scholar
  81. Yoo BJ, Spaete RR, Geballe AP, Selby M, Houghton M, Han JH (1992) 5’ end-dependent translation initiation of Hepatitis C viral RNA and the presence of putative positive and negative translational control elements within the 5’ untranslated region. Virology 191: 889–899PubMedCrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 1995

Authors and Affiliations

  • C. Wang
    • 1
    • 2
  • A. Siddiqui
    • 1
  1. 1.Department of Microbiology, Program in Molecular BiologyUniversity of Colorado Health Sciences CenterDenverUSA
  2. 2.Department of Cell BiologyHarvard Medical SchoolBostonUSA

Personalised recommendations