Skip to content
1887

Journal of General Virology header logo Journal of General Virology

Volume 85, Issue 7

Characterization of and phenotypes of poliovirus type 1 mutants with reduced viral protein synthesis activity No Access

Abstract

Sabin vaccine strains of poliovirus (PV) contain major attenuation determinants in the internal ribosomal entry site (IRES), an area that directs viral protein synthesis. To examine the effect of reduced viral protein synthesis on PV neurovirulence, spacer sequences, consisting of short open reading frames of different lengths, were introduced between the IRES and the initiation codon of viral polyprotein, resulting in PV mutants with reduced viral protein synthesis. These PV mutants had a viral protein synthesis activity 8·8–55 % of that of the parental Mahoney strain as measured in HeLa S3 cells. Only viruses with more than 28 % of the wild-type activity had intact spacer sequences following plaque purification. Mutants with 17 % or 21 % of the wild-type activity were unstable and a mutant with 8·8 % was lethal. The neurovirulence of PV mutants was evaluated in transgenic mice carrying the human PV receptor gene. In this test, mutants with more than 28 % of the wild-type activity remained neurovirulent, while a mutant with 17 % of wild-type activity exhibited a partially attenuated phenotype. This mutant stably replicated in the spinal cord; however, the stability was severely affected during the course of virus infection from the cerebrum to the spinal cord. These results suggest that reduced viral protein synthesis activity as measured in cultured cells (17–55 % of the wild-type activity) is not the main determinant of PV attenuation.

  • Received:
  • Accepted:
  • Published Online:
SGM
Loading

Article metrics loading...

/content/journal/jgv/10.1099/vir.0.19768-0
2004-07-01
2025-03-23
Loading full text...

Full text loading...

References

  1. Abe S., Ota Y., Koike S., Kurata T., Horie H., Nomura T., Hashizume S., Nomoto A. 1995; Neurovirulence test for oral live poliovaccines using poliovirus-sensitive transgenic mice. Virology 206:1075–1083 [CrossRef]
     [Google Scholar]
  2. Andino R., Rieckhof G. E., Baltimore D. 1990; A functional ribonucleoprotein complex forms around the 5′ end of poliovirus RNA. Cell 63:369–380 [CrossRef]
     [Google Scholar]
  3. Borman A. M., Deliat F. G., Kean K. M. 1994; Sequences within the poliovirus internal ribosome entry segment control viral RNA synthesis. EMBO J 13:3149–3157
     [Google Scholar]
  4. Bouchard M. J., Lam D. H., Racaniello V. R. 1995; Determinants of attenuation and temperature sensitivity in the type 1 poliovirus Sabin vaccine. J Virol 69:4972–4978
     [Google Scholar]
  5. Burns C. C., Richards O. C., Ehrenfeld E. 1992; Temperature-sensitive polioviruses containing mutations in RNA polymerase. Virology 189:568–582 [CrossRef]
     [Google Scholar]
  6. Cann A. J., Stanway G., Hughes P. J., Minor P. D., Evans D. M., Schild G. C., Almond J. W. 1984; Reversion to neurovirulence of the live-attenuated Sabin type 3 oral poliovirus vaccine. Nucleic Acids Res 12:7787–7792 [CrossRef]
     [Google Scholar]
  7. Chappell S. A., Edelman G. M., Mauro V. P. 2000; A 9-nt segment of a cellular mRNA can function as an internal ribosome entry site (IRES) and when present in linked multiple copies greatly enhances IRES activity. Proc Natl Acad Sci U S A 97:1536–1541 [CrossRef]
     [Google Scholar]
  8. Evans D. M., Dunn G., Minor P. D., Schild G. C., Cann A. J., Stanway G., Almond J. W., Currey K., Maizel J. V. 1985; Increased neurovirulence associated with a single nucleotide change in a noncoding region of the Sabin type 3 poliovaccine genome. Nature 314:548–550 [CrossRef]
     [Google Scholar]
  9. Garcia-Barrio M. T., Naranda T., Vazquez de Aldana C. R., Cuesta R., Hinnebusch A. G., Hershey J. W., Tamame M. 1995; GCD10, a translational repressor of GCN4, is the RNA-binding subunit of eukaryotic translation initiation factor-3. Genes Dev 9:1781–1796 [CrossRef]
     [Google Scholar]
  10. Gromeier M., Alexander L., Wimmer E. 1996; Internal ribosomal entry site substitution eliminates neurovirulence in intergeneric poliovirus recombinants. Proc Natl Acad Sci U S A 93:2370–2375 [CrossRef]
     [Google Scholar]
  11. Gutierrez A. L., Denova-Ocampo M., Racaniello V. R., del Angel R. M. 1997; Attenuating mutations in the poliovirus 5′ untranslated region alter its interaction with polypyrimidine tract-binding protein. J Virol 71:3826–3833
     [Google Scholar]
  12. Haller A. A., Stewart S. R., Semler B. L. 1996; Attenuation stem-loop lesions in the 5′ noncoding region of poliovirus RNA: neuronal cell-specific translation defects. J Virol 70:1467–1474
     [Google Scholar]
  13. Hellen C. U., Pestova T. V., Wimmer E. 1994; Effect of mutations downstream of the internal ribosome entry site on initiation of poliovirus protein synthesis. J Virol 68:6312–6322
     [Google Scholar]
  14. Herold J., Andino R. 2000; Poliovirus requires a precise 5′ end for efficient positive-strand RNA synthesis. J Virol 74:6394–6400 [CrossRef]
     [Google Scholar]
  15. Horie H., Koike S., Kurata T. 7 other authors 1994; Transgenic mice carrying the human poliovirus receptor: new animal models for study of poliovirus neurovirulence. J Virol 68:681–688
     [Google Scholar]
  16. Iizuka N., Kohara M., Hagino-Yamagishi K., Abe S., Komatsu T., Tago K., Arita M., Nomoto A. 1989; Construction of less neurovirulent polioviruses by introducing deletions into the 5′ noncoding sequence of the genome. J Virol 63:5354–5363
     [Google Scholar]
  17. Jang S. K., Krausslich H. G., Nicklin M. J., Duke G. M., Palmenberg A. C., Wimmer E. 1988; A segment of the 5′ nontranslated region of encephalomyocarditis virus RNA directs internal entry of ribosomes during in vitro translation. J Virol 62:2636–2643
     [Google Scholar]
  18. Johansen L. K., Morrow C. D. 2000; Inherent instability of poliovirus genomes containing two internal ribosome entry site (IRES) elements supports a role for the IRES in encapsidation. J Virol 74:8335–8342 [CrossRef]
     [Google Scholar]
  19. Karber G. 1931; Beitrag zur kollektiven Behandlung pharmakologischer Reihenversuche. Arch Exp Pathol Pharmakol 162:480–483 [CrossRef]
     [Google Scholar]
  20. Kawamura N., Kohara M., Abe S., Komatsu T., Tago K., Arita M., Nomoto A. 1989; Determinants in the 5′ noncoding region of poliovirus Sabin 1 RNA that influence the attenuation phenotype. J Virol 63:1302–1309
     [Google Scholar]
  21. Koike S., Taya C., Kurata T., Abe S., Ise I., Yonekawa H., Nomoto A. 1991; Transgenic mice susceptible to poliovirus. Proc Natl Acad Sci U S A 88:951–955 [CrossRef]
     [Google Scholar]
  22. Koike S., Taya C., Aoki J. 7 other authors 1994; Characterization of three different transgenic mouse lines that carry human poliovirus receptor gene – influence of the transgene expression on pathogenesis. Arch Virol 139:351–363 [CrossRef]
     [Google Scholar]
  23. Kozak M. 1986; Influences of mRNA secondary structure on initiation by eukaryotic ribosomes. Proc Natl Acad Sci U S A 83:2850–2854 [CrossRef]
     [Google Scholar]
  24. Kozak M. 1987; Effects of intercistronic length on the efficiency of reinitiation by eucaryotic ribosomes. Mol Cell Biol 7:3438–3445
     [Google Scholar]
  25. Kozak M. 1999; Initiation of translation in prokaryotes and eukaryotes. Gene 234:187–208 [CrossRef]
     [Google Scholar]
  26. Kuge S., Nomoto A. 1987; Construction of viable deletion and insertion mutants of the Sabin strain of type 1 poliovirus: function of the 5′ noncoding sequence in viral replication. J Virol 61:1478–1487
     [Google Scholar]
  27. Kuge S., Kawamura N., Nomoto A. 1989; Genetic variation occurring on the genome of an in vitro insertion mutant of poliovirus type 1. J Virol 63:1069–1075
     [Google Scholar]
  28. Li X., Lu H. H., Mueller S., Wimmer E. 2001; The C-terminal residues of poliovirus proteinase 2A(pro) are critical for viral RNA replication but not for cis- or trans-proteolytic cleavage. J Gen Virol 82:397–408
     [Google Scholar]
  29. Lu H. H., Alexander L., Wimmer E. 1995; Construction and genetic analysis of dicistronic polioviruses containing open reading frames for epitopes of human immunodeficiency virus type 1 gp120. J Virol 69:4797–4806
     [Google Scholar]
  30. Luukkonen B. G., Tan W., Schwartz S. 1995; Efficiency of reinitiation of translation on human immunodeficiency virus type 1 mRNAs is determined by the length of the upstream open reading frame and by intercistronic distance. J Virol 69:4086–4094
     [Google Scholar]
  31. Macadam A. J., Pollard S. R., Ferguson G., Dunn G., Skuce R., Almond J. W., Minor P. D. 1991; The 5′ noncoding region of the type 2 poliovirus vaccine strain contains determinants of attenuation and temperature sensitivity. Virology 181:451–458 [CrossRef]
     [Google Scholar]
  32. Macadam A. J., Pollard S. R., Ferguson G., Skuce R., Wood D., Almond J. W., Minor P. D. 1993; Genetic basis of attenuation of the Sabin type 2 vaccine strain of poliovirus in primates. Virology 192:18–26 [CrossRef]
     [Google Scholar]
  33. Macadam A. J., Stone D. M., Almond J. W., Minor P. D. 1994; The 5′ noncoding region and virulence of poliovirus vaccine strains. Trends Microbiol 2:449–454 [CrossRef]
     [Google Scholar]
  34. Malnou C. E., Poyry T. A., Jackson R. J., Kean K. M. 2002; Poliovirus internal ribosome entry segment structure alterations that specifically affect function in neuronal cells: molecular genetic analysis. J Virol 76:10617–10626 [CrossRef]
     [Google Scholar]
  35. Minor P. D. 1992; The molecular biology of poliovaccines. J Gen Virol 73:3065–3077 [CrossRef]
     [Google Scholar]
  36. Muzychenko A. R., Lipskaya G., Maslova S. V., Svitkin Y. V., Pilipenko E. V., Nottay B. K., Kew O. M., Agol V. I. 1991; Coupled mutations in the 5′-untranslated region of the Sabin poliovirus strains during in vivo passages: structural and functional implications. Virus Res 21:111–122 [CrossRef]
     [Google Scholar]
  37. Omata T., Kohara M., Kuge S. 7 other authors 1986; Genetic analysis of the attenuation phenotype of poliovirus type 1. J Virol 58:348–358
     [Google Scholar]
  38. Pelletier J., Sonenberg N. 1988; Internal initiation of translation of eukaryotic mRNA directed by a sequence derived from poliovirus RNA. Nature 334:320–325 [CrossRef]
     [Google Scholar]
  39. Pilipenko E. V., Viktorova E. G., Guest S. T., Agol V. I., Roos R. P. 2001; Cell-specific proteins regulate viral RNA translation and virus-induced disease. EMBO J 20:6899–6908 [CrossRef]
     [Google Scholar]
  40. Poyry T. A., Hentze M. W., Jackson R. J. 2001; Construction of regulatable picornavirus IRESes as a test of current models of the mechanism of internal translation initiation. RNA 7:647–660 [CrossRef]
     [Google Scholar]
  41. Ren R., Racaniello V. R. 1992; Human poliovirus receptor gene expression and poliovirus tissue tropism in transgenic mice. J Virol 66:296–304
     [Google Scholar]
  42. Rowe A., Burlison J., Macadam A. J., Minor P. D. 2001; Functional formation of domain V of the poliovirus noncoding region: significance of unpaired bases. Virology 289:45–53 [CrossRef]
     [Google Scholar]
  43. Sabin A. B. 1965; Oral poliovirus vaccine. History of its development and prospects for eradication of poliomyelitis. JAMA (J Am Med Assoc) 194:872–876 [CrossRef]
     [Google Scholar]
  44. Sambrook J., Russell D. W. 2001; In Molecular Cloning: a Laboratory Manual . , 3rd edn.pp. 13·19–13·25 Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
  45. Shiroki K., Kato H., Koike S., Odaka T., Nomoto A. 1993; Temperature-sensitive mouse cell factors for strand-specific initiation of poliovirus RNA synthesis. J Virol 67:3989–3996
     [Google Scholar]
  46. Shiroki K., Ishii T., Aoki T., Kobashi M., Ohka S., Nomoto A. 1995; A new cis-acting element for RNA replication within the 5′ noncoding region of poliovirus type 1 RNA. J Virol 69:6825–6832
     [Google Scholar]
  47. Shiroki K., Ishii T., Aoki T. 8 other authors 1997; Host range phenotype induced by mutations in the internal ribosomal entry site of poliovirus RNA. J Virol 71:1–8
     [Google Scholar]
  48. Slobodskaya O. R., Gmyl A. P., Maslova S. V., Tolskaya E. A., Viktorova E. G., Agol V. I. 1996; Poliovirus neurovirulence correlates with the presence of a cryptic AUG upstream of the initiator codon. Virology 221:141–150 [CrossRef]
     [Google Scholar]
  49. Stewart S. R., Semler B. L. 1999; Pyrimidine-rich region mutations compensate for a stem-loop V lesion in the 5′ noncoding region of poliovirus genomic RNA. Virology 264:385–397 [CrossRef]
     [Google Scholar]
  50. Svitkin Y. V., Maslova S. V., Agol V. I. 1985; The genomes of attenuated and virulent poliovirus strains differ in their in vitro translation efficiencies. Virology 147:243–252 [CrossRef]
     [Google Scholar]
  51. Svitkin Y. V., Cammack N., Minor P. D., Almond J. W. 1990; Translation deficiency of the Sabin type 3 poliovirus genome: association with an attenuating mutation C472–U. Virology 175:103–109 [CrossRef]
     [Google Scholar]
  52. Yoneyama T., Yoshida H., Shimizu H., Yoshii K., Nagata N., Kew O., Miyamura T. 2001; Neurovirulence of Sabin 1-derived polioviruses isolated from an immunodeficient patient with prolonged viral excretion. Dev Biol (Basel 105:93–98
     [Google Scholar]
  53. Zhao W. D., Wimmer E. 2001; Genetic analysis of a poliovirus/hepatitis C virus chimera: new structure for domain II of the internal ribosomal entry site of hepatitis C virus. J Virol 75:3719–3730 [CrossRef]
     [Google Scholar]
/content/journal/jgv/10.1099/vir.0.19768-0
Loading
Characterization of in vitro and in vivo phenotypes of poliovirus type 1 mutants with reduced viral protein synthesis activity
J. Gen. Virol. 85, 1933 (2004); https://doi.org/10.1099/vir.0.19768-0
/content/journal/jgv/10.1099/vir.0.19768-0
/content/journal/jgv/10.1099/vir.0.19768-0
Loading

Data & Media loading...

Supplements

Supplementary material 1

PDF

Most cited Most Cited RSS feed

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