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. 1986 May;83(10):3194–3198. doi: 10.1073/pnas.83.10.3194

Self-inactivating retroviral vectors designed for transfer of whole genes into mammalian cells.

S F Yu, T von Rüden, P W Kantoff, C Garber, M Seiberg, U Rüther, W F Anderson, E F Wagner, E Gilboa
PMCID: PMC323479  PMID: 3458176

Abstract

A retrovirus-derived vector called self-inactivating (SIN) vector was designed for the transduction of whole genes into mammalian cells. SIN vectors contain a deletion of 299 base pairs in the 3' long terminal repeat (LTR), which includes sequences encoding the enhancer and promoter functions. When viruses derived from such vectors were used to infect NIH 3T3 cells, the deletion was transferred to the 5' LTR, resulting in the transcriptional inactivation of the provirus in the infected cell. Introduction of a hybrid gene (human metallothionein-promoted c-fos) into cells via a SIN vector was not associated with rearrangements and led to the formation of an authentic mRNA transcript, which in some cases was induced by cadmium. SIN vectors should be particularly useful in gene transfer experiments designed to study the regulated expression of genes in mammalian cells. Absence of enhancer and promoter sequences in both LTRs of the integrated provirus should also minimize the possibility of activating cellular oncogenes and may provide a safer alternative to be used in human gene therapy.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Anderson W. F. Prospects for human gene therapy. Science. 1984 Oct 26;226(4673):401–409. doi: 10.1126/science.6093246. [DOI] [PubMed] [Google Scholar]
  2. Bandyopadhyay P. K., Temin H. M. Expression of complete chicken thymidine kinase gene inserted in a retrovirus vector. Mol Cell Biol. 1984 Apr;4(4):749–754. doi: 10.1128/mcb.4.4.749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Benz E. W., Jr, Wydro R. M., Nadal-Ginard B., Dina D. Moloney murine sarcoma proviral DNA is a transcriptional unit. Nature. 1980 Dec 25;288(5792):665–669. doi: 10.1038/288665a0. [DOI] [PubMed] [Google Scholar]
  4. Cepko C. L., Roberts B. E., Mulligan R. C. Construction and applications of a highly transmissible murine retrovirus shuttle vector. Cell. 1984 Jul;37(3):1053–1062. doi: 10.1016/0092-8674(84)90440-9. [DOI] [PubMed] [Google Scholar]
  5. Colbère-Garapin F., Horodniceanu F., Kourilsky P., Garapin A. C. A new dominant hybrid selective marker for higher eukaryotic cells. J Mol Biol. 1981 Jul 25;150(1):1–14. doi: 10.1016/0022-2836(81)90321-1. [DOI] [PubMed] [Google Scholar]
  6. Dick J. E., Magli M. C., Huszar D., Phillips R. A., Bernstein A. Introduction of a selectable gene into primitive stem cells capable of long-term reconstitution of the hemopoietic system of W/Wv mice. Cell. 1985 Aug;42(1):71–79. doi: 10.1016/s0092-8674(85)80102-1. [DOI] [PubMed] [Google Scholar]
  7. Eglitis M. A., Kantoff P., Gilboa E., Anderson W. F. Gene expression in mice after high efficiency retroviral-mediated gene transfer. Science. 1985 Dec 20;230(4732):1395–1398. doi: 10.1126/science.2999985. [DOI] [PubMed] [Google Scholar]
  8. Emerman M., Temin H. M. High-frequency deletion in recovered retrovirus vectors containing exogenous DNA with promoters. J Virol. 1984 Apr;50(1):42–49. doi: 10.1128/jvi.50.1.42-49.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Friedman R. L. Expression of human adenosine deaminase using a transmissable murine retrovirus vector system. Proc Natl Acad Sci U S A. 1985 Feb;82(3):703–707. doi: 10.1073/pnas.82.3.703. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Graves B. J., Eisenman R. N., McKnight S. L. Delineation of transcriptional control signals within the Moloney murine sarcoma virus long terminal repeat. Mol Cell Biol. 1985 Aug;5(8):1948–1958. doi: 10.1128/mcb.5.8.1948. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Hwang L. H., Gilboa E. Expression of genes introduced into cells by retroviral infection is more efficient than that of genes introduced into cells by DNA transfection. J Virol. 1984 May;50(2):417–424. doi: 10.1128/jvi.50.2.417-424.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jimenez A., Davies J. Expression of a transposable antibiotic resistance element in Saccharomyces. Nature. 1980 Oct 30;287(5785):869–871. doi: 10.1038/287869a0. [DOI] [PubMed] [Google Scholar]
  13. Jorgensen R. A., Rothstein S. J., Reznikoff W. S. A restriction enzyme cleavage map of Tn5 and location of a region encoding neomycin resistance. Mol Gen Genet. 1979;177(1):65–72. doi: 10.1007/BF00267254. [DOI] [PubMed] [Google Scholar]
  14. Joyner A. L., Bernstein A. Retrovirus transduction: segregation of the viral transforming function and the herpes simplex virus tk gene in infectious Friend spleen focus-forming virus thymidine kinase vectors. Mol Cell Biol. 1983 Dec;3(12):2191–2202. doi: 10.1128/mcb.3.12.2191. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Keller G., Paige C., Gilboa E., Wagner E. F. Expression of a foreign gene in myeloid and lymphoid cells derived from multipotent haematopoietic precursors. Nature. 1985 Nov 14;318(6042):149–154. doi: 10.1038/318149a0. [DOI] [PubMed] [Google Scholar]
  16. Lehrach H., Diamond D., Wozney J. M., Boedtker H. RNA molecular weight determinations by gel electrophoresis under denaturing conditions, a critical reexamination. Biochemistry. 1977 Oct 18;16(21):4743–4751. doi: 10.1021/bi00640a033. [DOI] [PubMed] [Google Scholar]
  17. Levinson B., Khoury G., Vande Woude G., Gruss P. Activation of SV40 genome by 72-base pair tandem repeats of Moloney sarcoma virus. Nature. 1982 Feb 18;295(5850):568–572. doi: 10.1038/295568a0. [DOI] [PubMed] [Google Scholar]
  18. Mann R., Mulligan R. C., Baltimore D. Construction of a retrovirus packaging mutant and its use to produce helper-free defective retrovirus. Cell. 1983 May;33(1):153–159. doi: 10.1016/0092-8674(83)90344-6. [DOI] [PubMed] [Google Scholar]
  19. Miller A. D., Eckner R. J., Jolly D. J., Friedmann T., Verma I. M. Expression of a retrovirus encoding human HPRT in mice. Science. 1984 Aug 10;225(4662):630–632. doi: 10.1126/science.6377498. [DOI] [PubMed] [Google Scholar]
  20. Miller A. D., Jolly D. J., Friedmann T., Verma I. M. A transmissible retrovirus expressing human hypoxanthine phosphoribosyltransferase (HPRT): gene transfer into cells obtained from humans deficient in HPRT. Proc Natl Acad Sci U S A. 1983 Aug;80(15):4709–4713. doi: 10.1073/pnas.80.15.4709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Miller A. D., Law M. F., Verma I. M. Generation of helper-free amphotropic retroviruses that transduce a dominant-acting, methotrexate-resistant dihydrofolate reductase gene. Mol Cell Biol. 1985 Mar;5(3):431–437. doi: 10.1128/mcb.5.3.431. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Rüther U., Wagner E. F., Müller R. Analysis of the differentiation-promoting potential of inducible c-fos genes introduced into embryonal carcinoma cells. EMBO J. 1985 Jul;4(7):1775–1781. doi: 10.1002/j.1460-2075.1985.tb03850.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Seiki M., Hattori S., Hirayama Y., Yoshida M. Human adult T-cell leukemia virus: complete nucleotide sequence of the provirus genome integrated in leukemia cell DNA. Proc Natl Acad Sci U S A. 1983 Jun;80(12):3618–3622. doi: 10.1073/pnas.80.12.3618. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Simonsen C. C., Levinson A. D. Isolation and expression of an altered mouse dihydrofolate reductase cDNA. Proc Natl Acad Sci U S A. 1983 May;80(9):2495–2499. doi: 10.1073/pnas.80.9.2495. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Southern E. M. Detection of specific sequences among DNA fragments separated by gel electrophoresis. J Mol Biol. 1975 Nov 5;98(3):503–517. doi: 10.1016/s0022-2836(75)80083-0. [DOI] [PubMed] [Google Scholar]
  26. Van Beveren C., van Straaten F., Curran T., Müller R., Verma I. M. Analysis of FBJ-MuSV provirus and c-fos (mouse) gene reveals that viral and cellular fos gene products have different carboxy termini. Cell. 1983 Apr;32(4):1241–1255. doi: 10.1016/0092-8674(83)90306-9. [DOI] [PubMed] [Google Scholar]
  27. Wagner E. F., Vanek M., Vennström B. Transfer of genes into embryonal carcinoma cells by retrovirus infection: efficient expression from an internal promoter. EMBO J. 1985 Mar;4(3):663–666. doi: 10.1002/j.1460-2075.1985.tb03680.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Wigler M., Silverstein S., Lee L. S., Pellicer A., Cheng Y. c., Axel R. Transfer of purified herpes virus thymidine kinase gene to cultured mouse cells. Cell. 1977 May;11(1):223–232. doi: 10.1016/0092-8674(77)90333-6. [DOI] [PubMed] [Google Scholar]
  29. Williams D. A., Lemischka I. R., Nathan D. G., Mulligan R. C. Introduction of new genetic material into pluripotent haematopoietic stem cells of the mouse. Nature. 1984 Aug 9;310(5977):476–480. doi: 10.1038/310476a0. [DOI] [PubMed] [Google Scholar]

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