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. 1985 Sep;5(9):2325–2331. doi: 10.1128/mcb.5.9.2325

Suppression of the hypomethylated Moloney leukemia virus genome in undifferentiated teratocarcinoma cells and inefficiency of transformation by a bacterial gene under control of the long terminal repeat.

O Niwa
PMCID: PMC366959  PMID: 3016527

Abstract

The Moloney leukemia virus (M-MuLV) genome was introduced into undifferentiated teratocarcinoma cells by transfection of a plasmid with the virus genome linked to pSV2neo, which carries a bacterial drug resistance gene, neo, or by cotransfection with pSV2neo. In the resulting cells, the M-MuLV genome remained hypomethylated, but its expression was suppressed in cells in an undifferentiated state. The pattern of DNA methylation of the viral genome remained unchanged when the cells were induced to differentiate into epithelial tissues. However, spontaneous M-MuLV expression was detected with differentiation of the cells. To determine to what extent the viral long terminal repeat (LTR) was responsible for this suppression in undifferentiated cells, I constructed plasmids in which neo was placed under the control of the promoter sequence of the dihydrofolate reductase gene or the M-MuLV LTR, and compared the biological activities of the plasmids in Ltk- cells and in undifferentiated teratocarcinoma cells. In Ltk- cells, these plasmids were highly efficient in making the cells resistant to selection by G418. However, in undifferentiated teratocarcinoma cells, the M-MuLV LTR promoted neo gene expression at only 10% of the expected efficiency, as compared with the expression of the neo gene under the control of the simian virus to or dihydrofolate reductase promoter. Thus, the mechanisms of gene regulation are not the same in undifferentiated and differentiated teratocarcinoma cells.

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Selected References

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

  1. Bacheler L., Fan H. Isolation of recombinant DNA clones carrying complete integrated proviruses of Moloney murine leukemia virus. J Virol. 1981 Jan;37(1):181–190. doi: 10.1128/jvi.37.1.181-190.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Chirgwin J. M., Przybyla A. E., MacDonald R. J., Rutter W. J. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochemistry. 1979 Nov 27;18(24):5294–5299. doi: 10.1021/bi00591a005. [DOI] [PubMed] [Google Scholar]
  3. Crouse G. F., Simonsen C. C., McEwan R. N., Schimke R. T. Structure of amplified normal and variant dihydrofolate reductase genes in mouse sarcoma S180 cells. J Biol Chem. 1982 Jul 10;257(13):7887–7897. [PubMed] [Google Scholar]
  4. Gautsch J. W. Embryonal carcinoma stem cells lack a function required for virus replication. Nature. 1980 May 8;285(5760):110–112. doi: 10.1038/285110a0. [DOI] [PubMed] [Google Scholar]
  5. Gautsch J. W., Wilson M. C. Delayed de novo methylation in teratocarcinoma suggests additional tissue-specific mechanisms for controlling gene expression. Nature. 1983 Jan 6;301(5895):32–37. doi: 10.1038/301032a0. [DOI] [PubMed] [Google Scholar]
  6. Graham F. L., van der Eb A. J. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology. 1973 Apr;52(2):456–467. doi: 10.1016/0042-6822(73)90341-3. [DOI] [PubMed] [Google Scholar]
  7. Groudine M., Eisenman R., Weintraub H. Chromatin structure of endogenous retroviral genes and activation by an inhibitor of DNA methylation. Nature. 1981 Jul 23;292(5821):311–317. doi: 10.1038/292311a0. [DOI] [PubMed] [Google Scholar]
  8. Harbers K., Schnieke A., Stuhlmann H., Jähner D., Jaenisch R. DNA methylation and gene expression: endogenous retroviral genome becomes infectious after molecular cloning. Proc Natl Acad Sci U S A. 1981 Dec;78(12):7609–7613. doi: 10.1073/pnas.78.12.7609. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hartley J. W., Rowe W. P. Clonal cells lines from a feral mouse embryo which lack host-range restrictions for murine leukemia viruses. Virology. 1975 May;65(1):128–134. doi: 10.1016/0042-6822(75)90013-6. [DOI] [PubMed] [Google Scholar]
  10. Hoffmann J. W., Steffen D., Gusella J., Tabin C., Bird S., Cowing D., Weinberg R. A. DNA methylation affecting the expression of murine leukemia proviruses. J Virol. 1982 Oct;44(1):144–157. doi: 10.1128/jvi.44.1.144-157.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Huang L. H., Wang R., Gama-Sosa M. A., Shenoy S., Ehrlich M. A protein from human placental nuclei binds preferentially to 5-methylcytosine-rich DNA. Nature. 1984 Mar 15;308(5956):293–295. doi: 10.1038/308293a0. [DOI] [PubMed] [Google Scholar]
  12. Huebner K., Linnenbach A., Weidner S., Glenn G., Croce C. M. Deoxyribonuclease I sensitivity of plasmid genomes in teratocarcinoma-derived stem and differentiated cells. Proc Natl Acad Sci U S A. 1981 Aug;78(8):5071–5075. doi: 10.1073/pnas.78.8.5071. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Linney E., Davis B., Overhauser J., Chao E., Fan H. Non-function of a Moloney murine leukaemia virus regulatory sequence in F9 embryonal carcinoma cells. 1984 Mar 29-Apr 4Nature. 308(5958):470–472. doi: 10.1038/308470a0. [DOI] [PubMed] [Google Scholar]
  14. Niwa O., Decléve A., Liberman M., Kaplan H. S. Adaptation of plaque assay methods to the in vitro quantitation of the radiation leukemia virus. J Virol. 1973 Jul;12(1):68–73. doi: 10.1128/jvi.12.1.68-73.1973. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Niwa O., Sugahara T. 5-Azacytidine induction of mouse endogenous type C virus and suppression of DNA methylation. Proc Natl Acad Sci U S A. 1981 Oct;78(10):6290–6294. doi: 10.1073/pnas.78.10.6290. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Niwa O., Yokota Y., Ishida H., Sugahara T. Independent mechanisms involved in suppression of the Moloney leukemia virus genome during differentiation of murine teratocarcinoma cells. Cell. 1983 Apr;32(4):1105–1113. doi: 10.1016/0092-8674(83)90294-5. [DOI] [PubMed] [Google Scholar]
  17. Segal S., Levine A. J., Khoury G. Evidence for non-spliced SV40 RNA in undifferentiated murine teratocarcinoma stem cells. Nature. 1979 Jul 26;280(5720):335–338. doi: 10.1038/280335a0. [DOI] [PubMed] [Google Scholar]
  18. Simon D., Stuhlmann H., Jähner D., Wagner H., Werner E., Jaenisch R. Retrovirus genomes methylated by mammalian but not bacterial methylase are non-infectious. Nature. 1983 Jul 21;304(5923):275–277. doi: 10.1038/304275a0. [DOI] [PubMed] [Google Scholar]
  19. 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]
  20. Southern P. J., Berg P. Transformation of mammalian cells to antibiotic resistance with a bacterial gene under control of the SV40 early region promoter. J Mol Appl Genet. 1982;1(4):327–341. [PubMed] [Google Scholar]
  21. Speers W. C., Gautsch J. W., Dixon F. J. Silent infection of murine embryonal carcinoma cells by Moloney murine leukemia virus. Virology. 1980 Aug;105(1):241–244. doi: 10.1016/0042-6822(80)90171-3. [DOI] [PubMed] [Google Scholar]
  22. Stalder J., Larsen A., Engel J. D., Dolan M., Groudine M., Weintraub H. Tissue-specific DNA cleavages in the globin chromatin domain introduced by DNAase I. Cell. 1980 Jun;20(2):451–460. doi: 10.1016/0092-8674(80)90631-5. [DOI] [PubMed] [Google Scholar]
  23. Stewart C. L., Stuhlmann H., Jähner D., Jaenisch R. De novo methylation, expression, and infectivity of retroviral genomes introduced into embryonal carcinoma cells. Proc Natl Acad Sci U S A. 1982 Jul;79(13):4098–4102. doi: 10.1073/pnas.79.13.4098. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Stuhlmann H., Jähner D., Jaenisch R. Infectivity and methylation of retroviral genomes is correlated with expression in the animal. Cell. 1981 Oct;26(2 Pt 2):221–232. doi: 10.1016/0092-8674(81)90305-6. [DOI] [PubMed] [Google Scholar]
  25. Tan K. B., Huebner K., Croce C. M. Chromosomal proteins of mouse teratocarcinoma cells. J Cell Physiol Suppl. 1982;2:51–59. doi: 10.1002/jcp.1041130510. [DOI] [PubMed] [Google Scholar]
  26. Thomas P. S. Hybridization of denatured RNA and small DNA fragments transferred to nitrocellulose. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5201–5205. doi: 10.1073/pnas.77.9.5201. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Weintraub H., Larsen A., Groudine M. Alpha-Globin-gene switching during the development of chicken embryos: expression and chromosome structure. Cell. 1981 May;24(2):333–344. doi: 10.1016/0092-8674(81)90323-8. [DOI] [PubMed] [Google Scholar]
  28. Wigler M., Sweet R., Sim G. K., Wold B., Pellicer A., Lacy E., Maniatis T., Silverstein S., Axel R. Transformation of mammalian cells with genes from procaryotes and eucaryotes. Cell. 1979 Apr;16(4):777–785. doi: 10.1016/0092-8674(79)90093-x. [DOI] [PubMed] [Google Scholar]

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