Abstract
Infection by murine retroviruses in embryonic carcinoma (EC) and embryonic stem cells is highly restricted. The transcriptional unit of the Moloney murine leukemic virus (MoMuLV) long terminal repeat (LTR) is inactive in EC and embryonic stem cells in association with increased proviral methylation. In this study, expression in F9 EC cells was achieved from novel retroviral vectors containing three modifications in the MoMuLV-based retroviral vector: presence of the myeloproliferative sarcoma virus LTR, substitution of the primer binding site, and either deletion of a negative control region at the 5' end of the LTR or insertion of a demethylating sequence. We conclude that inhibition of expression from the MoMuLV LTR in EC cells is mediated through the additive effects of multiple cis-acting elements affecting the state of methylation of the provirus.
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- Akgün E., Ziegler M., Grez M. Determinants of retrovirus gene expression in embryonal carcinoma cells. J Virol. 1991 Jan;65(1):382–388. doi: 10.1128/jvi.65.1.382-388.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bowtell D. D., Cory S., Johnson G. R., Gonda T. J. Comparison of expression in hemopoietic cells by retroviral vectors carrying two genes. J Virol. 1988 Jul;62(7):2464–2473. doi: 10.1128/jvi.62.7.2464-2473.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Bowtell D. D., Johnson G. R., Kelso A., Cory S. Expression of genes transferred to haemopoietic stem cells by recombinant retroviruses. Mol Biol Med. 1987 Aug;4(4):229–250. [PubMed] [Google Scholar]
- Boyes J., Bird A. DNA methylation inhibits transcription indirectly via a methyl-CpG binding protein. Cell. 1991 Mar 22;64(6):1123–1134. doi: 10.1016/0092-8674(91)90267-3. [DOI] [PubMed] [Google Scholar]
- Cedar H. DNA methylation and gene activity. Cell. 1988 Apr 8;53(1):3–4. doi: 10.1016/0092-8674(88)90479-5. [DOI] [PubMed] [Google Scholar]
- Cepko C. L., Ryder E. F., Austin C. P., Walsh C., Fekete D. M. Lineage analysis using retrovirus vectors. Methods Enzymol. 1993;225:933–960. doi: 10.1016/0076-6879(93)25059-b. [DOI] [PubMed] [Google Scholar]
- Challita P. M., Kohn D. B. Lack of expression from a retroviral vector after transduction of murine hematopoietic stem cells is associated with methylation in vivo. Proc Natl Acad Sci U S A. 1994 Mar 29;91(7):2567–2571. doi: 10.1073/pnas.91.7.2567. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Chirigos M. A., Scott D., Turner W., Perk K. Biological, pathological and physical characterization of a possible variant of a murine sarcoma virus (Moloney). Int J Cancer. 1968 Mar 15;3(2):223–227. doi: 10.1002/ijc.2910030207. [DOI] [PubMed] [Google Scholar]
- Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
- Colicelli J., Goff S. P. Isolation of a recombinant murine leukemia virus utilizing a new primer tRNA. J Virol. 1986 Jan;57(1):37–45. doi: 10.1128/jvi.57.1.37-45.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Feinberg A. P., Vogelstein B. "A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity". Addendum. Anal Biochem. 1984 Feb;137(1):266–267. doi: 10.1016/0003-2697(84)90381-6. [DOI] [PubMed] [Google Scholar]
- Feuer G., Taketo M., Hanecak R. C., Fan H. Two blocks in Moloney murine leukemia virus expression in undifferentiated F9 embryonal carcinoma cells as determined by transient expression assays. J Virol. 1989 May;63(5):2317–2324. doi: 10.1128/jvi.63.5.2317-2324.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Flanagan J. R., Becker K. G., Ennist D. L., Gleason S. L., Driggers P. H., Levi B. Z., Appella E., Ozato K. Cloning of a negative transcription factor that binds to the upstream conserved region of Moloney murine leukemia virus. Mol Cell Biol. 1992 Jan;12(1):38–44. doi: 10.1128/mcb.12.1.38. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Flanagan J. R., Krieg A. M., Max E. E., Khan A. S. Negative control region at the 5' end of murine leukemia virus long terminal repeats. Mol Cell Biol. 1989 Feb;9(2):739–746. doi: 10.1128/mcb.9.2.739. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Gorman C. M., Rigby P. W., Lane D. P. Negative regulation of viral enhancers in undifferentiated embryonic stem cells. Cell. 1985 Sep;42(2):519–526. doi: 10.1016/0092-8674(85)90109-6. [DOI] [PubMed] [Google Scholar]
- Grez M., Zörnig M., Nowock J., Ziegler M. A single point mutation activates the Moloney murine leukemia virus long terminal repeat in embryonal stem cells. J Virol. 1991 Sep;65(9):4691–4698. doi: 10.1128/jvi.65.9.4691-4698.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Guild B. C., Finer M. H., Housman D. E., Mulligan R. C. Development of retrovirus vectors useful for expressing genes in cultured murine embryonal cells and hematopoietic cells in vivo. J Virol. 1988 Oct;62(10):3795–3801. doi: 10.1128/jvi.62.10.3795-3801.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Hilberg F., Stocking C., Ostertag W., Grez M. Functional analysis of a retroviral host-range mutant: altered long terminal repeat sequences allow expression in embryonal carcinoma cells. Proc Natl Acad Sci U S A. 1987 Aug;84(15):5232–5236. doi: 10.1073/pnas.84.15.5232. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Jaenisch R. Transgenic animals. Science. 1988 Jun 10;240(4858):1468–1474. doi: 10.1126/science.3287623. [DOI] [PubMed] [Google Scholar]
- Keller G., Kennedy M., Papayannopoulou T., Wiles M. V. Hematopoietic commitment during embryonic stem cell differentiation in culture. Mol Cell Biol. 1993 Jan;13(1):473–486. doi: 10.1128/mcb.13.1.473. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Kempler G., Freitag B., Berwin B., Nanassy O., Barklis E. Characterization of the Moloney murine leukemia virus stem cell-specific repressor binding site. Virology. 1993 Apr;193(2):690–699. doi: 10.1006/viro.1993.1177. [DOI] [PubMed] [Google Scholar]
- 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]
- Loh T. P., Sievert L. L., Scott R. W. Evidence for a stem cell-specific repressor of Moloney murine leukemia virus expression in embryonal carcinoma cells. Mol Cell Biol. 1990 Aug;10(8):4045–4057. doi: 10.1128/mcb.10.8.4045. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Loh T. P., Sievert L. L., Scott R. W. Proviral sequences that restrict retroviral expression in mouse embryonal carcinoma cells. Mol Cell Biol. 1987 Oct;7(10):3775–3784. doi: 10.1128/mcb.7.10.3775. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Markowitz D., Goff S., Bank A. A safe packaging line for gene transfer: separating viral genes on two different plasmids. J Virol. 1988 Apr;62(4):1120–1124. doi: 10.1128/jvi.62.4.1120-1124.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- 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]
- Ostertag W., Vehmeyer K., Fagg B., Pragnell I. B., Paetz W., Le Bousse M. C., Smadja-Joffe F., Klein B., Jasmin C., Eisen H. Myeloproliferative virus, a cloned murine sarcoma virus with spleen focus-forming properties in adult mice. J Virol. 1980 Feb;33(2):573–582. doi: 10.1128/jvi.33.2.573-582.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Petersen R., Kempler G., Barklis E. A stem cell-specific silencer in the primer-binding site of a retrovirus. Mol Cell Biol. 1991 Mar;11(3):1214–1221. doi: 10.1128/mcb.11.3.1214. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Prince V. E., Rigby P. W. Derivatives of Moloney murine sarcoma virus capable of being transcribed in embryonal carcinoma stem cells have gained a functional Sp1 binding site. J Virol. 1991 Apr;65(4):1803–1811. doi: 10.1128/jvi.65.4.1803-1811.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Seliger B., Kollek R., Stocking C., Franz T., Ostertag W. Viral transfer, transcription, and rescue of a selectable myeloproliferative sarcoma virus in embryonal cell lines: expression of the mos oncogene. Mol Cell Biol. 1986 Jan;6(1):286–293. doi: 10.1128/mcb.6.1.286. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Soriano P., Friedrich G., Lawinger P. Promoter interactions in retrovirus vectors introduced into fibroblasts and embryonic stem cells. J Virol. 1991 May;65(5):2314–2319. doi: 10.1128/jvi.65.5.2314-2319.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 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]
- Stocking C., Kollek R., Bergholz U., Ostertag W. Long terminal repeat sequences impart hematopoietic transformation properties to the myeloproliferative sarcoma virus. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5746–5750. doi: 10.1073/pnas.82.17.5746. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Stocking C., Kollek R., Bergholz U., Ostertag W. Point mutations in the U3 region of the long terminal repeat of Moloney murine leukemia virus determine disease specificity of the myeloproliferative sarcoma virus. Virology. 1986 Aug;153(1):145–149. doi: 10.1016/0042-6822(86)90015-2. [DOI] [PubMed] [Google Scholar]
- Szyf M., Tanigawa G., McCarthy P. L., Jr A DNA signal from the Thy-1 gene defines de novo methylation patterns in embryonic stem cells. Mol Cell Biol. 1990 Aug;10(8):4396–4400. doi: 10.1128/mcb.10.8.4396. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Taketo M., Shaffer D. J. Deletions in a recombinant retrovirus genome associated with its expression in embryonal carcinoma cells. J Virol. 1989 Oct;63(10):4431–4433. doi: 10.1128/jvi.63.10.4431-4433.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tsukiyama T., Niwa O., Yokoro K. Mechanism of suppression of the long terminal repeat of Moloney leukemia virus in mouse embryonal carcinoma cells. Mol Cell Biol. 1989 Nov;9(11):4670–4676. doi: 10.1128/mcb.9.11.4670. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Tsukiyama T., Ueda H., Hirose S., Niwa O. Embryonal long terminal repeat-binding protein is a murine homolog of FTZ-F1, a member of the steroid receptor superfamily. Mol Cell Biol. 1992 Mar;12(3):1286–1291. doi: 10.1128/mcb.12.3.1286. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Weiher H., Barklis E., Ostertag W., Jaenisch R. Two distinct sequence elements mediate retroviral gene expression in embryonal carcinoma cells. J Virol. 1987 Sep;61(9):2742–2746. doi: 10.1128/jvi.61.9.2742-2746.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Wiles M. V. Embryonic stem cell differentiation in vitro. Methods Enzymol. 1993;225:900–918. doi: 10.1016/0076-6879(93)25057-9. [DOI] [PubMed] [Google Scholar]