Advertisement

Retroviral Vectors as Insertional Mutagens

  • Joop Gäken
  • Farzin Farzaneh
Part of the Methods in Molecular Biology book series (MIMB, volume 8)

Abstract

A critical step in the life-cycle of retroviruses is the integration of the double-stranded DNA copy of their RNA genome into the genome of the host cell (1). Although the provirus DNA sequences flanking the site of integration are precisely determined and characteristic of each virus (2), the vast majority of the integrations are the product of nonhomologous recomination events, resulting in the pseudorandom integration of the provirus into the host-cell genome (3,4). Retroviruses can therefore act as agents of insertional mutagenesis. The insertion of the provirus into the genome could, in principle, result in either gene activation or gene inactivation. The insertional inactivation would be the result of provirus integration within the coding or regulatory sequences of the gene of interest, thus disrupting the expression of a functional gene product (5, 6, 7, 8, 9, 10, 11, 12, 13). Insertional activation could be the product of the integration of viral-promoter enhancer elements, contained within the long terminal repeat (LTR) sequences in the vicinity of a silent gene, resulting in the increased transcription of that gene (14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26). In addition to these direct as-acting effects, there can be indirect trans-regulatory effects resulting from the presence of the viral genome within the cell, but irrespective of its position of integration. This could be the product of genes or other regulatory elements encoded by the virus. The position-independent effect(s) of retrovirus integration would be easy to identify; they would be present in all cells or in a vastly larger number of cells than would be compatible with low-frequency integrations into specific genomic domains.

Keywords

Long Terminal Repeat Insertional Mutagenesis Embryonal Carcinoma Cell Packaging Cell Long Terminal Repeat Sequence 
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.

References

  1. 1.
    Varmus, H. (1988) Retroviruses. Science 240, 1427–1435.PubMedCrossRefGoogle Scholar
  2. 2.
    Varmus, H. and Swanstrom, R. (1982) Replication of retroviruses, in RNA Tumour Viruses (Weiss, R., Teich, N., Varmus, H., and Coffin, J., eds.), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY, pp. 369–512.Google Scholar
  3. 3.
    Hughes, S. H., Shank, P. R., Spector, D. H., Rung, H. G., Bishop, J. M., Varmus, H. E., Vogt, P. K., and Breitman, M. L. (1978) Proviruses of avian sarcoma virus are terminally redundant, coextensive with unintegrated linear DNA and integrated at many sites. Cell 15, 1397–1410.PubMedCrossRefGoogle Scholar
  4. 4.
    Shimotohno, K. and Temin, H. M. (1980) No apparent nucleotide sequence specificity in cellular DNA juxtaposed to retrovirus proviruses. Proc. Natl. Acad. Sci. USA 77, 7357–7361.PubMedCrossRefGoogle Scholar
  5. 5.
    Jenkins, N. A., Copeland, N. G., Taylor, B. A., and Lee, B. K. (1981) Dilute (d) coat colour mutation of DBA/2J mice is associated with the site of integration of an ecotropic MuLV genome. Nature 293, 370–374.PubMedCrossRefGoogle Scholar
  6. 6.
    Copeland, N. G., Jenkins, N. A., and Lee, B. K. (1983) Association of the lethal yellow (Ay) coat color mutation with the ecotropic murine leukemia virus genome. Proc. Natl. Acad. Sci. USA 80, 247–249.PubMedCrossRefGoogle Scholar
  7. 7.
    Copeland, N. G., Hutchison, K. W., and Jenkins, N. A. (1983) Excision of the DBA ecotropic provirus m dilute coat/color revertants of mice occurs by homologous recombination involving the viral LTRs. Cell 33, 379–387.PubMedCrossRefGoogle Scholar
  8. 8.
    Schnieke, A., Harbers, K., and Jaenisch, R. (1983) Embryonic lethal mutation in mice induced by retrovirus insertion into the alpha 1 (I) collagen gene. Nature 304, 315–320.PubMedCrossRefGoogle Scholar
  9. 9.
    Jaenisch, R., Harbers, K., Schnieke, A., Lohler, J., Chumakov, I., Jahner, D., Grotkopp, D., and Hoffmann, E. (1983) Germ line integration of Moloney murine leukemia virus at the Movl 3 locus leads to recessive lethal mutation and early embryonic death. Cell 32, 209–216.PubMedCrossRefGoogle Scholar
  10. 10.
    Wolf, D. and Rotter, V. (1984) Inactivation of p53 gene expression by an insertion of Moloney murine leukemia virus-like DNA sequence. Mol. Cell Biol. 4,1402–1410.PubMedGoogle Scholar
  11. 11.
    Frankel, W., Potter, T. A., Rosenberg, N., Lenz, J., and Rajan, T. V. (1985) Retroviral insertional mutagenesis of a target allele in a heterozygous murine cell line. Proc. Natl. Acad. Sci. USA 82,6600–6604.PubMedCrossRefGoogle Scholar
  12. 12.
    King, W., Patel, M. D., Lobel, L. I., Goff, S. P., and Chi Nguyen-Huu, M. (1985) Insertion mutagenesis of embryonal carcinoma cells by retroviruses. Science 228, 554–558.PubMedCrossRefGoogle Scholar
  13. 13.
    Kuehn, M. R., Bradley, A., Robertson, E. J., and Evans, M. J. (1987) A potential animal model for Lesch-Nyhan syndrome through introduction of HPRT mutations into mice. Nature 326, 295–298.PubMedCrossRefGoogle Scholar
  14. 14.
    Hayward, W. S., Neel, B. G., and Astrin, S. M. (1981) Activation of a cellular one gene by promoter insertion in ALV-induced lymphoid leukosis. Nature 290, 475–480.PubMedCrossRefGoogle Scholar
  15. 15.
    Varmus, H. E., Quintrell, N., and Oritz, S. (1981) Retroviruses as mutagens: Insertion and excision of a nontransforming provirus alters expression of a resident transforming provirus. Cell 25, 23–36.PubMedCrossRefGoogle Scholar
  16. 16.
    Nusse, R. and Varmus, H. E. (1982) Many tumors induced by the mouse mammary tumor virus contain a provirus integrated in the same region of the host cell genome. Cell 31, 99–109.PubMedCrossRefGoogle Scholar
  17. 17.
    Fung, Y. K. T., Lewis, W. G., Crittenden, L B., and Rung, H. J (1983) Activation of the cellular oncogene c-erbB by LTR insertion: Molecular basis for induction of erythroblastosis by avian leukosis virus. Cell 33, 357–368.PubMedCrossRefGoogle Scholar
  18. 18.
    Peters, G., Brookes, S., Smith, R., and Dickson, C. (1983) Tumorigenesis by mouse mammary tumour virus: Evidence for a common region for provirus integration in mammary tumours. Cell 33, 369–377.PubMedCrossRefGoogle Scholar
  19. 19.
    Tsichlis, N., Strauss, P. G., and Hu, L. F. (1983) A common region for proviral DNA integration in MoMuLV-induced rat thymic lymphomas. Nature 302, 445–449PubMedCrossRefGoogle Scholar
  20. 20.
    Wagner, E. F., Covarrubias, L., Stewart, T. A., and Mintz, B. (1983) Lethalities in mice homozygous for human growth hormone gene sequences integrated in the germ line. Cell 35, 647–655.PubMedCrossRefGoogle Scholar
  21. 21.
    Dickson, C., Smith, R., Brookes, S., and Peters, G. (1984) Tumorigenesis by mouse mammary tumor virus: Proviral activation of a cellular gene in the common integration region int-2. Cell 37, 529–536.PubMedCrossRefGoogle Scholar
  22. 22.
    Steffen, D. (1984) Proviruses are adjacent to c-myc in some murine leukemia virus-induced lymphomas. Proc. Natl. Acad. Sci. USA 81, 2097–2101.PubMedCrossRefGoogle Scholar
  23. 23.
    Shen-Ong, G. L. C., Potter, M., Mushinski, J. F., Lavu, S., and Reddy, E. P. (1984) Activation of c-myb locus by viral insertional mutagenesis in plasmacytoid lymphosarcomas. Science 226, 1077–1080.PubMedCrossRefGoogle Scholar
  24. 24.
    Lemay, G. and Jolicoeur, P. (1984) Rearrangement of a DNA sequence homologous to a cell-virus junction fragment in several Moloney murine leukemia virus-induced rat thymomas. Proc. Natl. Acad. Sci. USA 81, 38–42.PubMedCrossRefGoogle Scholar
  25. 25.
    Cuypers, H. T., Selten, G., Quint, W., Zijlstra, M., Maandag, R. E., Boelens, W., van Wezenbeek, P., Melief, C., and Berns, A. (1984) Murine leukemia virus induced T-cell lymphomagenesis: Integration of proviruses in a distinct chromosomal region. Cell 37 141–150.PubMedCrossRefGoogle Scholar
  26. 26.
    Stocking, C., Löliger, C., Kawai, M., Suciu, S., Gough, N., and Ostertag, W. (1988) Identification of genes involved in growth autonomy of haematopoietic cells by analysis of factor-independent mutants. Cell 53, 869–879.PubMedCrossRefGoogle Scholar
  27. 27.
    Scherdin, U., Rhodes, K., and Breindl, M (1990) Transcriptionally active genome regions are preferred targets for retrovirus integration. J. Virol. 64, 907–912.PubMedGoogle Scholar
  28. 28.
    Kozak, C. A. (1985) Susceptibility of wild mouse cells to exogenous infection with xenotropic leukemia viruses: Control by a single dominant locus on chromosome 1. J. Virol. 55, 690–695.PubMedGoogle Scholar
  29. 29.
    Sarrna, P. S., Cheong, M. P., and Hartley, J. W. (1967) A viral influence test for mouse leukemia viruses. Virology 33, 180–184.CrossRefGoogle Scholar
  30. 80.
    Stewart, C. L., Stuhlmann, H., Jahner, D., and Jaenisch, R. (1982) De novo methylation, expression, and infectivity of retroviral genomes introduced into embryonal carcinoma cells. Proc. Natl. Acad. Sci. USA 79, 4098–4102.PubMedCrossRefGoogle Scholar
  31. 31.
    Hooper, M. L. (1985) Mammalian Cell Genetics. Wiley, New York.Google Scholar
  32. 32.
    Reik, W., Weiher, H., and Jaenisch, R. (1984) Replication-competent Moloney murine leukemia virus carrying a bacterial suppressor tRNA gene: Selective cloning of proviral and flanking host sequences. Proc. Natl. Acad. Sci. USA 82, 1141–1145.CrossRefGoogle Scholar
  33. 33.
    Lobel, L. I., Patel, M., King, W., Nguyen-Huu, M. C, and Goff, S. P. (1985) Construction and recovery of viable retroviral genomes carrying a bacterial suppressor transfer RNA gene. Science 228, 329–332.PubMedCrossRefGoogle Scholar
  34. 34.
    Saiki, R. K., Scharf, S., Faloona, F., Mullis, K. B., Horn, G T., Erlich, H. A., and Amheim, N. (1985) Enzymatic amplification of beta-globin genomic sequences and restriction site analysis for diagnosis of Sickle cell anaemia. Science 230, 1350–1354.PubMedCrossRefGoogle Scholar
  35. 35.
    Silver, J. and Keerikatte, V. (1989) Novel use of polymerase chain reaction to amplify cellular DNA adjacent to an integrated provirus. J. Virol. 63, 1925–1928.Google Scholar
  36. 36.
    Wong, C., Dowling, C. E., Saiki, K, Higuchi, R. G., Erlich, H. A., and Kazazian, H H. (1987) Characterization of β-thalassaemia mutations using direct genomic sequencing of amplified single copy DNA. Nature 330, 384–386.PubMedCrossRefGoogle Scholar
  37. 37.
    Miller, A. D. and Buttimore, C. (1986) Redesign of retrovirus packaging cell lines to avoid recombination leading to helper virus production. Mol. Cell Biol. 6, 2895–2902.PubMedGoogle Scholar
  38. 38.
    Markowitz, D., Goff, S., and Bank, A. (1988) A safe packaging line for gene transfer: separating viral genes on two different plasmids. J Virol. 62, 1120–1124.PubMedGoogle Scholar
  39. 39.
    Stacey, A., Arbuthnott, C., Kollek, R., Coggins, L., and Ostertag, W. (1984) Comparison of myeloproliferative sarcoma virus with Moloney murine sarcoma virus variants by nucleotide sequencing and heteroduplex analysis.J. Virol. 50, 725–732.PubMedGoogle Scholar
  40. 40.
    Markowitz, D., Goff, S., and Bank, A. (1988) Construction and use of a safe and efficient amphotropic packaging cell line. Virology 167, 400–406.PubMedGoogle Scholar
  41. 41.
    Danos, O. and Mulligan, R. C. (1988) Safe and efficient generation of recombinant retrovirus with amphotropic and ecotropic host ranges. Proc. Natl. Acad. Sci. USA 85, 6460–6464.PubMedCrossRefGoogle Scholar
  42. 42.
    Albritton, L. M., Tseng, L., Scadden, D., and Cunningham, J. M. (1989) A putative murine ecotropic retrovirus receptor gene encodes a multiple membrane-spanning protein and confers susceptibility to virus infection. Cell 57, 659–666.PubMedCrossRefGoogle Scholar
  43. 43.
    Scadden, D. T., Fuller, B., and Cunningham, J. M. (1990) Human cells infected with retrovirus vectors acquire an endogenous murine provirus. J Virol. 64, 424–427.PubMedGoogle Scholar

Copyright information

© The Humana Press Inc., Clifton, NJ 1991

Authors and Affiliations

  • Joop Gäken
    • 1
  • Farzin Farzaneh
    • 1
  1. 1.Molecular Genetics UnitKing’s College School of Medicine and DentistryLondonUK

Personalised recommendations