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A Significantly Improved Semliki Forest Virus Expression System Based on Translation Enhancer Segments from the Viral Capsid Gene

Bio/Technology volume 12, pages 1127–1131 (1994)Cite this article

Abstract

We recently described a system for heterologous gene expression in a variety of mammalian cell types that is based on an efficiently replicating Semliki Forest virus (SFV) variant in which an RNA encoding a foreign protein replaces the RNA that normally encodes the viruses' structural polyprotein. Although expression levels are sufficiently high for many purposes, in general they are only 10% of the level of the polyprotein in a wild type SFV infection. Here we show that the first 102 bases of the viral capsid gene function as a translational enhancer, and that SFV vectors incorporating this RNA increase heterologous protein synthesis to the level of wild type polyprotein.

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References

  1. Liljeström, P. and Garoff, H. 1991. A new generation of animal cell expression vectors based on the Semliki Forest virus replicon. Bio/Technology 9: 1356–1361.

    Article  Google Scholar 

  2. Berglund, P., Sjöberg, M., Garoff, H., Atkins, G., Sheahan, B. and Liljeström, P. 1993. Semliki Forest virus expression system: production of conditionally infectious recombinant particles. Bio/Technology 11: 916–920.

    CAS  Google Scholar 

  3. Xiong,C., Levis, R., Shen, P., Schlesinger, S., Rice, C.M. and Huang, H.V. 1989. Sindbis virus: An efficient, broad host range vector for gene expression in animal cells. Science 243: 1181–1191.

    Article  Google Scholar 

  4. Ding, M. and Schlesinger, M.J. 1989. Evidence that Sindbis virus nsP2 is an autoprotease which processes the virus nonstructural polyprotein. Virology 171: 280–284.

    Article  CAS  PubMed  Google Scholar 

  5. Hardy, W.R. and Strauss, J.H. 1989. Processing the nonstructural polyproteins of Sindbis virus: nonstructural proteinase is in the C-terminal half of nsP2 and functions both in cis and in trans. J. Virol. 63: 4653–4664.

    CAS  PubMed  PubMed Central  Google Scholar 

  6. Strauss, E.G. and Strauss, J.H. 1986. Structure and replication of the alpha virus genome, p. 35–90. In: The Togaviridae and Flaviviridae, S.S. Schlesinger and M.J. Schlesinger (Eds. ). Plenum Press, New York.

    Chapter  Google Scholar 

  7. Aliperti, G. and Schlesinger, M.J. 1978. Evidence for an autoprotease activity of Sindbis virus capsid protein. Virology 90: 366–369

    Article  CAS  PubMed  Google Scholar 

  8. Choi, H.-K., Tong, L., Minor, W., Dumas, P., Boege, U., Rossmann, M.G. and Wengler, G. .1991. Structure of Sindbis virus core protein reveals a chymotrypsin-like serine proteinase and the organization of the virion. Nature 354: 37–43.

    Article  CAS  PubMed  Google Scholar 

  9. Hahn, C.S. and Strauss, J.H. 1990. Site-directed mutagenesis of the proposed catalytic amino acids of the Sindbis virus capsid protein autoprotease. J. Virol. 64: 3069–3073.

    CAS  PubMed  PubMed Central  Google Scholar 

  10. Melançon, P. and Garoff, H. 1987. Processing of the Semliki Forest virus structural polyprotein: Role of the capsid protease. J. Virol. 61: 1301–1309.

    PubMed  PubMed Central  Google Scholar 

  11. Garoff, H., Huylebroeck, D., Robinson, A., Tillman, U. and Liljeström, P. 1990. The signal sequence of the p62 protein of Semliki Forest virus is involved in initiation but not in completing chain translocation. J. Cell Biol. 111: 867–876.

    Article  CAS  PubMed  Google Scholar 

  12. Garoff, H., Simons, K. and Dobberstein, B. B.1978. Assembly of Semliki Forest virus membrane glycoproteins in the membrane of the endoplasmic reticulum in vitro. J. Mol. Biol. 124: 587–600.

    Article  CAS  PubMed  Google Scholar 

  13. Hashimoto, K., Erdei, S., Keränen, S., Saraste, J. and Kääriäinen, L. 1981. Evidence for a separate signal sequence for the carboxy-terminal envelope glycoprotein E1 of Semliki Forest virus. J. Virol. 38: 34–40.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Liljeström, P. and Garoff, H. 1991. Internally located cleavable signal sequences direct the formation of Semliki Forest virus membrane proteins from a polyprotein precursor. J. Virol. 65: 147–154.

    PubMed  PubMed Central  Google Scholar 

  15. Melançon, P. and Garoff, H. 1986. Reinitiation of translocation in the Semliki Forest virus structural polyprotein: Identification of the signal for the E1 glycoprotein. EMBO J. 5: 1551–1560.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Forsell, K., Suomalainen, M. and Garoff, H. 1994. Structure/function relation of the RNA binding domain of the Semliki Forest virus capsid protein. In preparation

    Google Scholar 

  17. de Curtis, I. and Simons, K. 1988. Dissection of Semliki Forest virus glycoprotein delivery from the trans-Golgi network to the cell surface in permeabilized BHK cells. Proc. Natl. Acad. Sci. USA 85: 8052–8056.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Barth, B.-U., Wahlberg, J.M. and Garoff, H. 1994. The oligomerization reaction of the Semliki Forest virus membrane protein subunits. In preparation.

    Google Scholar 

  19. Yoshinaka, Y., Katoh, I., Copeland, T. and Oroszlan, S. 1985. Murine leukemia virus protease is encoded by the gag-pol gene and is synthesized through suppression of an amber termination codon. Proc. Natl. Acad. Sci. USA 82: 1618–1622.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Wills, N.M., Gestland, R.F. and Atkins, J.F. 1991. Evidence that a down stream pseudoknot is required for translational read-through of the Moloney murine leukemia virus gag stop codon. Proc. Natl. Acad. Sci. USA 88: 6691–6995.

    Article  Google Scholar 

  21. Garry, R.F. 1994. Sindbis virus-induced inhibition of protein synthesis is partially reversed by medium containing an elevated potassium concentration. J. Gen. Virol. 75: 411–415.

    Article  CAS  PubMed  Google Scholar 

  22. Suomalainen, M. and Garoff, H. 1994. Incorporation of homologous and heterologous proteins into the envelop of Moloney leukemia virus. J. Virol. 68: 4879–4889.

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Liljeström, P., Lusa, S., Huylebroeck, D. and Garoff, H. 1991. In vitro mutagenesis of a full-length cDNA clone of Semliki Forest virus: the 6,000-molecular-weight membrane protein modulates virus release. J. Virol. 65: 4107–4113.

    PubMed  PubMed Central  Google Scholar 

  24. Suomalainen, M., Liljeström, P. and Garoff, H. 1992. Spike protein nucleocapsid interactions drive the budding of alphaviruses. J. Virol. 66: 4737–4747.

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Peränen, J., Takkinen, K., Kalkkinen, N. and Kääriäinen, L. 1988. Semliki Forest virus-specific non-structural protein nsP3 is a phosphoprotein. J. Gen. Virol. 69: 2165–2178.

    Article  PubMed  Google Scholar 

  26. Suomalainen, M., Baron, M. and Garoff, H. 1990. The E2 signal sequence of Rubella virus remains part of the capsid protein and confers membrane association in vitro. J. Virol. 64: 5500–5509.

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Sambrook, J., Fritsch, E.F. and Maniatis, T. 1989. Molecular Cloning. A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.

    Google Scholar 

  28. Boere, W.A.M., Harmsen, T., Vinje, J., Benaissa-Trouw, B.J., Kraaijeeveld, C.A. and Snippe, H. 1984. Identification of distinct antigenic determinants on Semliki Forest virus by using monoclonal antibodies with different antiviral activities. J. Virol. 52: 575–582.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Wahlberg, J.M., Boere, W.A. and Garoff, H. 1989. The heterodimeric association between the membrane proteins of Semliki Forest virus changes its sensitivity to mildly acidic pH during virus maturation. J. Virol. 63: 4991–4997.

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Maizel, J.V. Jr. 1971. Polyacrylamide gel electrophoresis of viral proteins. Methods Virol. 5: 179–246.

    Article  Google Scholar 

  31. Sanes, J., Rubenstein, J.L.R. and Nicolas, J.-F. 1986. Use of recombinant retrovirus to study post-implantation cell lineage in mouse embryos. EMBO J. 5: 3133–3142.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Authors and Affiliations

  1. Center for Biotechnology, Novum, S-141 57, Huddinge, Sweden

    E. Mathilda Sjöberg, Maarit Suomalainen & Henrik Garoff

Authors
  1. E. Mathilda Sjöberg
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  2. Maarit Suomalainen
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  3. Henrik Garoff
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Correspondence to E. Mathilda Sjöberg.

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Sjöberg, E., Suomalainen, M. & Garoff, H. A Significantly Improved Semliki Forest Virus Expression System Based on Translation Enhancer Segments from the Viral Capsid Gene. Nat Biotechnol 12, 1127–1131 (1994). https://doi.org/10.1038/nbt1194-1127

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