Skip to main content
NCBI home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1994 Apr 25;22(8):1484–1488. doi: 10.1093/nar/22.8.1484

Retrotransposition of a marked Drosophila line-like I element in cells in culture.

S Jensen 1, L Cavarec 1, O Dhellin 1, T Heidmann 1
PMCID: PMC308009  PMID: 8190641

Abstract

We have marked a Drosophila transposable element--the LINE-like I element--with an intron-containing indicator gene inserted in place of a large deletion in the I element second ORF encompassing the reverse transcriptase domain, and this marked element was placed downstream to a potent actin promoter. An expression vector for the I element ORFs was also constructed, under the same heterologous promoter. The indicator gene contains a lacZ reporter gene the expression of which is conditioned by retrotransposition of the marked element, thus allowing detection of transposition events by testing for either beta-galactosidase expression or occurrence of spliced DNA molecules. The marked I element was introduced into Drosophila melanogaster cells in culture by transfection. Spliced DNA copies of the marked element and specifically stained beta-galactosidase-expressing cells were detected only upon co-transfection with the I expression vector, thus indicating that an ORF2-deleted element can be complemented in trans for transposition. This simple assay for retrotransposition in Drosophila cells in culture provides a tool for the rapid analysis of the mechanism of I transposition in its cis and trans sequence requirements.

Full text

PDF
1484

Images in this article

1486Image
on p.1486
1486Image
on p.1486

Selected References

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

  1. Bonnerot C., Rocancourt D., Briand P., Grimber G., Nicolas J. F. A beta-galactosidase hybrid protein targeted to nuclei as a marker for developmental studies. Proc Natl Acad Sci U S A. 1987 Oct;84(19):6795–6799. doi: 10.1073/pnas.84.19.6795. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bucheton A. I transposable elements and I-R hybrid dysgenesis in Drosophila. Trends Genet. 1990 Jan;6(1):16–21. doi: 10.1016/0168-9525(90)90044-7. [DOI] [PubMed] [Google Scholar]
  3. Bucheton A., Paro R., Sang H. M., Pelisson A., Finnegan D. J. The molecular basis of I-R hybrid dysgenesis in Drosophila melanogaster: identification, cloning, and properties of the I factor. Cell. 1984 Aug;38(1):153–163. doi: 10.1016/0092-8674(84)90536-1. [DOI] [PubMed] [Google Scholar]
  4. Deragon J. M., Sinnett D., Labuda D. Reverse transcriptase activity from human embryonal carcinoma cells NTera2D1. EMBO J. 1990 Oct;9(10):3363–3368. doi: 10.1002/j.1460-2075.1990.tb07537.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Echalier G., Ohanessian A. In vitro culture of Drosophila melanogaster embryonic cells. In Vitro. 1970 Nov-Dec;6(3):162–172. doi: 10.1007/BF02617759. [DOI] [PubMed] [Google Scholar]
  6. Fawcett D. H., Lister C. K., Kellett E., Finnegan D. J. Transposable elements controlling I-R hybrid dysgenesis in D. melanogaster are similar to mammalian LINEs. Cell. 1986 Dec 26;47(6):1007–1015. doi: 10.1016/0092-8674(86)90815-9. [DOI] [PubMed] [Google Scholar]
  7. Gabriel A., Boeke J. D. Reverse transcriptase encoded by a retrotransposon from the trypanosomatid Crithidia fasciculata. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9794–9798. doi: 10.1073/pnas.88.21.9794. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Han K., Levine M. S., Manley J. L. Synergistic activation and repression of transcription by Drosophila homeobox proteins. Cell. 1989 Feb 24;56(4):573–583. doi: 10.1016/0092-8674(89)90580-1. [DOI] [PubMed] [Google Scholar]
  9. Heidmann O., Heidmann T. Retrotransposition of a mouse IAP sequence tagged with an indicator gene. Cell. 1991 Jan 11;64(1):159–170. doi: 10.1016/0092-8674(91)90217-m. [DOI] [PubMed] [Google Scholar]
  10. Heidmann T., Heidmann O., Nicolas J. F. An indicator gene to demonstrate intracellular transposition of defective retroviruses. Proc Natl Acad Sci U S A. 1988 Apr;85(7):2219–2223. doi: 10.1073/pnas.85.7.2219. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Ivanov V. A., Melnikov A. A., Siunov A. V., Fodor I. I., Ilyin Y. V. Authentic reverse transcriptase is coded by jockey, a mobile Drosophila element related to mammalian LINEs. EMBO J. 1991 Sep;10(9):2489–2495. doi: 10.1002/j.1460-2075.1991.tb07788.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Jensen S., Heidmann T. An indicator gene for detection of germline retrotransposition in transgenic Drosophila demonstrates RNA-mediated transposition of the LINE I element. EMBO J. 1991 Jul;10(7):1927–1937. doi: 10.1002/j.1460-2075.1991.tb07719.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Martin S. L., Branciforte D. Synchronous expression of LINE-1 RNA and protein in mouse embryonal carcinoma cells. Mol Cell Biol. 1993 Sep;13(9):5383–5392. doi: 10.1128/mcb.13.9.5383. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Martin S. L. LINEs. Curr Opin Genet Dev. 1991 Dec;1(4):505–508. doi: 10.1016/s0959-437x(05)80199-6. [DOI] [PubMed] [Google Scholar]
  15. Martin S. L. Ribonucleoprotein particles with LINE-1 RNA in mouse embryonal carcinoma cells. Mol Cell Biol. 1991 Sep;11(9):4804–4807. doi: 10.1128/mcb.11.9.4804. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Mathias S. L., Scott A. F., Kazazian H. H., Jr, Boeke J. D., Gabriel A. Reverse transcriptase encoded by a human transposable element. Science. 1991 Dec 20;254(5039):1808–1810. doi: 10.1126/science.1722352. [DOI] [PubMed] [Google Scholar]
  17. McLean C., Bucheton A., Finnegan D. J. The 5' untranslated region of the I factor, a long interspersed nuclear element-like retrotransposon of Drosophila melanogaster, contains an internal promoter and sequences that regulate expression. Mol Cell Biol. 1993 Feb;13(2):1042–1050. doi: 10.1128/mcb.13.2.1042. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Mizrokhi L. J., Georgieva S. G., Ilyin Y. V. jockey, a mobile Drosophila element similar to mammalian LINEs, is transcribed from the internal promoter by RNA polymerase II. Cell. 1988 Aug 26;54(5):685–691. doi: 10.1016/s0092-8674(88)80013-8. [DOI] [PubMed] [Google Scholar]
  19. Pritchard M. A., Dura J. M., Pélisson A., Bucheton A., Finnegan D. J. A cloned I-factor is fully functional in Drosophila melanogaster. Mol Gen Genet. 1988 Nov;214(3):533–540. doi: 10.1007/BF00330491. [DOI] [PubMed] [Google Scholar]
  20. Pélisson A., Finnegan D. J., Bucheton A. Evidence for retrotransposition of the I factor, a LINE element of Drosophila melanogaster. Proc Natl Acad Sci U S A. 1991 Jun 1;88(11):4907–4910. doi: 10.1073/pnas.88.11.4907. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Sanes J. R., Rubenstein J. L., Nicolas J. F. Use of a recombinant retrovirus to study post-implantation cell lineage in mouse embryos. EMBO J. 1986 Dec 1;5(12):3133–3142. doi: 10.1002/j.1460-2075.1986.tb04620.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Segal-Bendirdjian E., Heidmann T. Evidence for a reverse transcription intermediate for a marked line transposon in tumoral rat cells. Biochem Biophys Res Commun. 1991 Dec 16;181(2):863–870. doi: 10.1016/0006-291x(91)91270-m. [DOI] [PubMed] [Google Scholar]
  23. Steller H., Pirrotta V. A transposable P vector that confers selectable G418 resistance to Drosophila larvae. EMBO J. 1985 Jan;4(1):167–171. doi: 10.1002/j.1460-2075.1985.tb02332.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Swergold G. D. Identification, characterization, and cell specificity of a human LINE-1 promoter. Mol Cell Biol. 1990 Dec;10(12):6718–6729. doi: 10.1128/mcb.10.12.6718. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Tchenio T., Heidmann T. Defective retroviruses can disperse in the human genome by intracellular transposition. J Virol. 1991 Apr;65(4):2113–2118. doi: 10.1128/jvi.65.4.2113-2118.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Tchenio T., Heidmann T. High-frequency intracellular transposition of a defective mammalian provirus detected by an in situ colorimetric assay. J Virol. 1992 Mar;66(3):1571–1578. doi: 10.1128/jvi.66.3.1571-1578.1992. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Tchénio T., Segal-Bendirdjian E., Heidmann T. Generation of processed pseudogenes in murine cells. EMBO J. 1993 Apr;12(4):1487–1497. doi: 10.1002/j.1460-2075.1993.tb05792.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Xiong Y., Eickbush T. H. Origin and evolution of retroelements based upon their reverse transcriptase sequences. EMBO J. 1990 Oct;9(10):3353–3362. doi: 10.1002/j.1460-2075.1990.tb07536.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES