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. 1997 Sep;71(9):7128–7131. doi: 10.1128/jvi.71.9.7128-7131.1997

A conditional self-inactivating retrovirus vector that uses a tetracycline-responsive expression system.

J J Hwang 1, L Li 1, W F Anderson 1
PMCID: PMC192013  PMID: 9261449

Abstract

We developed a novel conditional self-inactivating (C-SIN) vector, TL-SN, by replacement of the enhancer-promoter of the 3' long terminal repeat of Moloney murine leukemia virus with a synthetic tetracycline operator-cytomegalovirus promoter (tetP) from the tetracycline-responsive expression system (TRES). The other component of the TRES, a chimeric transactivator (tTA), was stably incorporated into PA317 amphotropic packaging cells, thus generating the packaging cell line PA317-tTA. C-SIN amphotropic G418-resistant virus particles were generated with a titer of 2 x 10(5) CFU/ml within 2 days of transinfection of PA317-tTA cells with TL-SN ecotropic virus particles. This titer was approximately 2 log units higher than that obtained by transinfection of parental PA317 cells and was due to the high level of viral transcripts originating from the tetP promoter at the 5' end of the transduced vector in the presence of tTA. Our C-SIN vector has the potential for use in human gene therapy since it incorporates the advantages of previous SIN vectors in having weak tetP promoter activity (in the absence of tTA in target cells) while at the same time achieving high viral titers with PA317-tTA packaging cells.

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Selected References

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  1. Anderson W. F. Human gene therapy. Science. 1992 May 8;256(5058):808–813. doi: 10.1126/science.1589762. [DOI] [PubMed] [Google Scholar]
  2. Bishop J. M. Cellular oncogenes and retroviruses. Annu Rev Biochem. 1983;52:301–354. doi: 10.1146/annurev.bi.52.070183.001505. [DOI] [PubMed] [Google Scholar]
  3. Dougherty J. P., Temin H. M. A promoterless retroviral vector indicates that there are sequences in U3 required for 3' RNA processing. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1197–1201. doi: 10.1073/pnas.84.5.1197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Eglitis M. A., Anderson W. F. Retroviral vectors for introduction of genes into mammalian cells. Biotechniques. 1988 Jul-Aug;6(7):608–614. [PubMed] [Google Scholar]
  5. Emerman M., Temin H. M. Comparison of promoter suppression in avian and murine retrovirus vectors. Nucleic Acids Res. 1986 Dec 9;14(23):9381–9396. doi: 10.1093/nar/14.23.9381. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Emerman M., Temin H. M. Genes with promoters in retrovirus vectors can be independently suppressed by an epigenetic mechanism. Cell. 1984 Dec;39(3 Pt 2):449–467. [PubMed] [Google Scholar]
  7. Gossen M., Bujard H. Tight control of gene expression in mammalian cells by tetracycline-responsive promoters. Proc Natl Acad Sci U S A. 1992 Jun 15;89(12):5547–5551. doi: 10.1073/pnas.89.12.5547. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Hawley R. G., Covarrubias L., Hawley T., Mintz B. Handicapped retroviral vectors efficiently transduce foreign genes into hematopoietic stem cells. Proc Natl Acad Sci U S A. 1987 Apr;84(8):2406–2410. doi: 10.1073/pnas.84.8.2406. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hawley R. G., Sabourin L. A., Hawley T. S. An improved retroviral vector for gene transfer into undifferentiated cells. Nucleic Acids Res. 1989 May 25;17(10):4001–4001. doi: 10.1093/nar/17.10.4001. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Hofmann A., Nolan G. P., Blau H. M. Rapid retroviral delivery of tetracycline-inducible genes in a single autoregulatory cassette. Proc Natl Acad Sci U S A. 1996 May 28;93(11):5185–5190. doi: 10.1073/pnas.93.11.5185. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hoshimaru M., Ray J., Sah D. W., Gage F. H. Differentiation of the immortalized adult neuronal progenitor cell line HC2S2 into neurons by regulatable suppression of the v-myc oncogene. Proc Natl Acad Sci U S A. 1996 Feb 20;93(4):1518–1523. doi: 10.1073/pnas.93.4.1518. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Hwang J. J., Scuric Z., Anderson W. F. Novel retroviral vector transferring a suicide gene and a selectable marker gene with enhanced gene expression by using a tetracycline-responsive expression system. J Virol. 1996 Nov;70(11):8138–8141. doi: 10.1128/jvi.70.11.8138-8141.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Iida A., Chen S. T., Friedmann T., Yee J. K. Inducible gene expression by retrovirus-mediated transfer of a modified tetracycline-regulated system. J Virol. 1996 Sep;70(9):6054–6059. doi: 10.1128/jvi.70.9.6054-6059.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Linial M., Groudine M. Transcription of three c-myc exons is enhanced in chicken bursal lymphoma cell lines. Proc Natl Acad Sci U S A. 1985 Jan;82(1):53–57. doi: 10.1073/pnas.82.1.53. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Miller A. D., Buttimore C. Redesign of retrovirus packaging cell lines to avoid recombination leading to helper virus production. Mol Cell Biol. 1986 Aug;6(8):2895–2902. doi: 10.1128/mcb.6.8.2895. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Miller A. D. Human gene therapy comes of age. Nature. 1992 Jun 11;357(6378):455–460. doi: 10.1038/357455a0. [DOI] [PubMed] [Google Scholar]
  17. Miller A. D., Rosman G. J. Improved retroviral vectors for gene transfer and expression. Biotechniques. 1989 Oct;7(9):980-2, 984-6, 989-90. [PMC free article] [PubMed] [Google Scholar]
  18. Olson P., Nelson S., Dornburg R. Improved self-inactivating retroviral vectors derived from spleen necrosis virus. J Virol. 1994 Nov;68(11):7060–7066. doi: 10.1128/jvi.68.11.7060-7066.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Osborne W. R. Retrovirus-mediated gene expression in mammalian cells. Curr Opin Biotechnol. 1991 Oct;2(5):708–712. doi: 10.1016/0958-1669(91)90039-8. [DOI] [PubMed] [Google Scholar]
  20. Paulus W., Baur I., Boyce F. M., Breakefield X. O., Reeves S. A. Self-contained, tetracycline-regulated retroviral vector system for gene delivery to mammalian cells. J Virol. 1996 Jan;70(1):62–67. doi: 10.1128/jvi.70.1.62-67.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Payne G. S., Bishop J. M., Varmus H. E. Multiple arrangements of viral DNA and an activated host oncogene in bursal lymphomas. Nature. 1982 Jan 21;295(5846):209–214. doi: 10.1038/295209a0. [DOI] [PubMed] [Google Scholar]
  22. Salmons B., Günzburg W. H. Targeting of retroviral vectors for gene therapy. Hum Gene Ther. 1993 Apr;4(2):129–141. doi: 10.1089/hum.1993.4.2-129. [DOI] [PubMed] [Google Scholar]
  23. Temin H. M. Safety considerations in somatic gene therapy of human disease with retrovirus vectors. Hum Gene Ther. 1990 Summer;1(2):111–123. doi: 10.1089/hum.1990.1.2-111. [DOI] [PubMed] [Google Scholar]
  24. Westaway D., Payne G., Varmus H. E. Proviral deletions and oncogene base-substitutions in insertionally mutagenized c-myc alleles may contribute to the progression of avian bursal tumors. Proc Natl Acad Sci U S A. 1984 Feb;81(3):843–847. doi: 10.1073/pnas.81.3.843. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Xu L., Yee J. K., Wolff J. A., Friedmann T. Factors affecting long-term stability of Moloney murine leukemia virus-based vectors. Virology. 1989 Aug;171(2):331–341. doi: 10.1016/0042-6822(89)90600-4. [DOI] [PubMed] [Google Scholar]
  26. Yarranton G. T. Inducible vectors for expression in mammalian cells. Curr Opin Biotechnol. 1992 Oct;3(5):506–511. doi: 10.1016/0958-1669(92)90078-w. [DOI] [PubMed] [Google Scholar]
  27. Yee J. K., Moores J. C., Jolly D. J., Wolff J. A., Respess J. G., Friedmann T. Gene expression from transcriptionally disabled retroviral vectors. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5197–5201. doi: 10.1073/pnas.84.15.5197. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Yu S. F., von Rüden T., Kantoff P. W., Garber C., Seiberg M., Rüther U., Anderson W. F., Wagner E. F., Gilboa E. Self-inactivating retroviral vectors designed for transfer of whole genes into mammalian cells. Proc Natl Acad Sci U S A. 1986 May;83(10):3194–3198. doi: 10.1073/pnas.83.10.3194. [DOI] [PMC free article] [PubMed] [Google Scholar]

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