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. 1986 Feb;83(3):812–816. doi: 10.1073/pnas.83.3.812

Expression of the oncogene of avian reticuloendotheliosis virus in Escherichia coli and identification of the transforming protein in reticuloendotheliosis virus T-transformed cells.

N K Herzog, H R Bose Jr
PMCID: PMC322955  PMID: 3003751

Abstract

The genome of reticuloendotheliosis virus T (REV-T) includes a unique oncogene v-rel, which is transcribed in low amounts into a 3.0-kilobase subgenomic mRNA in REV-T-transformed lymphoid cells. To identify the v-rel protein, REV-T DNA sequences were cloned into bacterial plasmid vectors designed to achieve expression of foreign DNA sequences in Escherichia coli. Portions of the v-rel gene were joined to the 5' segment of the trpE gene. Upon induction of trpE with indoleacrylic acid, large amounts of trpE-v-rel fusion proteins were produced by the bacteria carrying these recombinant plasmids. Two trpE-v-rel fusion proteins were synthesized in E. coli, which collectively represent three-quarters of the predicted v-rel protein. Polyclonal antisera were generated to trpE-v-red fusion proteins. These antisera were used in immunoblotting experiments to identify a 57-kDa v-rel protein in REV-T-transformed lymphoid cells lines and REV-T-infected chicken embryo fibroblast cultures. The v-rel gene expressed in E. coli under lac control was found to produce a 56-kDa protein. Although REV-T-transformed and Marek disease virus-transformed lymphoid cells contain c-rel mRNA transcripts, a c-rel protein could not be detected with antisera directed against v-rel fusion proteins.

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Selected References

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

  1. Adler H. I., Fisher W. D., Cohen A., Hardigree A. A. MINIATURE escherichia coli CELLS DEFICIENT IN DNA. Proc Natl Acad Sci U S A. 1967 Feb;57(2):321–326. doi: 10.1073/pnas.57.2.321. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Beug H., Müller H., Grieser S., Doederlein G., Graf T. Hematopoietic cells transformed in vitro by REVT avian reticuloendotheliosis virus express characteristics of very immature lymphoid cells. Virology. 1981 Dec;115(2):295–309. doi: 10.1016/0042-6822(81)90112-4. [DOI] [PubMed] [Google Scholar]
  3. Beug H., von Kirchbach A., Döderlein G., Conscience J. F., Graf T. Chicken hematopoietic cells transformed by seven strains of defective avian leukemia viruses display three distinct phenotypes of differentiation. Cell. 1979 Oct;18(2):375–390. doi: 10.1016/0092-8674(79)90057-6. [DOI] [PubMed] [Google Scholar]
  4. Breitman M. L., Lai M. M., Vogt P. K. The genomic RNA of avian reticuloendotheliosis virus REV. Virology. 1980 Jan 30;100(2):450–461. doi: 10.1016/0042-6822(80)90535-8. [DOI] [PubMed] [Google Scholar]
  5. Burnette W. N. "Western blotting": electrophoretic transfer of proteins from sodium dodecyl sulfate--polyacrylamide gels to unmodified nitrocellulose and radiographic detection with antibody and radioiodinated protein A. Anal Biochem. 1981 Apr;112(2):195–203. doi: 10.1016/0003-2697(81)90281-5. [DOI] [PubMed] [Google Scholar]
  6. Chen I. S., Mak T. W., O'Rear J. J., Temin H. M. Characterization of reticuloendotheliosis virus strain T DNA and isolation of a novel variant of reticuloendotheliosis virus strain T by molecular cloning. J Virol. 1981 Dec;40(3):800–811. doi: 10.1128/jvi.40.3.800-811.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Chen I. S., Wilhelmsen K. C., Temin H. M. Structure and expression of c-rel, the cellular homolog to the oncogene of reticuloendotheliosis virus strain T. J Virol. 1983 Jan;45(1):104–113. doi: 10.1128/jvi.45.1.104-113.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Franklin R. B., Kang C. Y., Min-Min Wan K., Bose H. R., Jr Transformation of chick embryo fibroblasts by reticuloendotheliosis virus. Virology. 1977 Dec;83(2):313–321. doi: 10.1016/0042-6822(77)90176-3. [DOI] [PubMed] [Google Scholar]
  9. Franklin R. B., Maldonado R. L., Bose H. R. Isolation and characterization of reticuloendotheliosis virus transformed bone marrow cells. Intervirology. 1974;3(5-6):342–352. doi: 10.1159/000149771. [DOI] [PubMed] [Google Scholar]
  10. Goeddel D. V., Kleid D. G., Bolivar F., Heyneker H. L., Yansura D. G., Crea R., Hirose T., Kraszewski A., Itakura K., Riggs A. D. Expression in Escherichia coli of chemically synthesized genes for human insulin. Proc Natl Acad Sci U S A. 1979 Jan;76(1):106–110. doi: 10.1073/pnas.76.1.106. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Gonda M. A., Rice N. R., Gilden R. V. Avian reticuloendotheliosis virus: characterization of the high-molecular-weight viral RNA in transforming and helper virus populations. J Virol. 1980 Jun;34(3):743–751. doi: 10.1128/jvi.34.3.743-751.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Graf T., Beug H. Avian leukemia viruses: interaction with their target cells in vivo and in vitro. Biochim Biophys Acta. 1978 Nov 17;516(3):269–299. doi: 10.1016/0304-419x(78)90011-2. [DOI] [PubMed] [Google Scholar]
  13. Hancock K., Tsang V. C. India ink staining of proteins on nitrocellulose paper. Anal Biochem. 1983 Aug;133(1):157–162. doi: 10.1016/0003-2697(83)90237-3. [DOI] [PubMed] [Google Scholar]
  14. Hoelzer J. D., Franklin R. B., Bose H. R., Jr Transformation by reticuloendotheliosis virus: development of a focus assay and isolation of a nontransforming virus. Virology. 1979 Feb;93(1):20–30. doi: 10.1016/0042-6822(79)90272-1. [DOI] [PubMed] [Google Scholar]
  15. Hoelzer J. D., Lewis R. B., Wasmuth C. R., Bose H. R., Jr Hematopoietic cell transformation by reticuloendotheliosis virus: characterization of the genetic defect. Virology. 1980 Jan 30;100(2):462–474. doi: 10.1016/0042-6822(80)90536-x. [DOI] [PubMed] [Google Scholar]
  16. Kleid D. G., Yansura D., Small B., Dowbenko D., Moore D. M., Grubman M. J., McKercher P. D., Morgan D. O., Robertson B. H., Bachrach H. L. Cloned viral protein vaccine for foot-and-mouth disease: responses in cattle and swine. Science. 1981 Dec 4;214(4525):1125–1129. doi: 10.1126/science.6272395. [DOI] [PubMed] [Google Scholar]
  17. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  18. Lederberg E. M., Cohen S. N. Transformation of Salmonella typhimurium by plasmid deoxyribonucleic acid. J Bacteriol. 1974 Sep;119(3):1072–1074. doi: 10.1128/jb.119.3.1072-1074.1974. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Levy S. B. Physical and functional characteristics of R-factor deoxyribonucleic acid segregated into Escherichia coli minicells. J Bacteriol. 1971 Oct;108(1):300–308. doi: 10.1128/jb.108.1.300-308.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Lewis R. B., McClure J., Rup B., Niesel D. W., Garry R. F., Hoelzer J. D., Nazerian K., Bose H. R., Jr Avian reticuloendotheliosis virus: identification of the hematopoietic target cell for transformation. Cell. 1981 Aug;25(2):421–431. doi: 10.1016/0092-8674(81)90060-x. [DOI] [PubMed] [Google Scholar]
  21. Markwell M. A. A new solid-state reagent to iodinate proteins. I. Conditions for the efficient labeling of antiserum. Anal Biochem. 1982 Sep 15;125(2):427–432. doi: 10.1016/0003-2697(82)90025-2. [DOI] [PubMed] [Google Scholar]
  22. Meagher R. B., Tait R. C., Betlach M., Boyer H. W. Protein expression in E. coli minicells by recombinant plasmids. Cell. 1977 Mar;10(3):521–536. doi: 10.1016/0092-8674(77)90039-3. [DOI] [PubMed] [Google Scholar]
  23. Norrander J., Kempe T., Messing J. Construction of improved M13 vectors using oligodeoxynucleotide-directed mutagenesis. Gene. 1983 Dec;26(1):101–106. doi: 10.1016/0378-1119(83)90040-9. [DOI] [PubMed] [Google Scholar]
  24. Roozen K. J., Fenwick R. G., Jr, Curtiss R., 3rd Synthesis of ribonucleic acid and protein in plasmid-containing minicells of Escherichia coli K-12. J Bacteriol. 1971 Jul;107(1):21–33. doi: 10.1128/jb.107.1.21-33.1971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Shibuya T., Chen I., Howatson A., Mak T. W. Morphological, immunological, and biochemical analyses of chicken spleen cells transformed in vitro by reticuloendotheliosis virus strain T. Cancer Res. 1982 Jul;42(7):2722–2728. [PubMed] [Google Scholar]
  26. Spindler K. R., Rosser D. S., Berk A. J. Analysis of adenovirus transforming proteins from early regions 1A and 1B with antisera to inducible fusion antigens produced in Escherichia coli. J Virol. 1984 Jan;49(1):132–141. doi: 10.1128/jvi.49.1.132-141.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Stephens R. M., Rice N. R., Hiebsch R. R., Bose H. R., Jr, Gilden R. V. Nucleotide sequence of v-rel: the oncogene of reticuloendotheliosis virus. Proc Natl Acad Sci U S A. 1983 Oct;80(20):6229–6233. doi: 10.1073/pnas.80.20.6229. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Van Beveren C., Galleshaw J. A., Jonas V., Berns A. J., Doolittle R. F., Donoghue D. J., Verma I. M. Nucleotide sequence and formation of the transforming gene of a mouse sarcoma virus. Nature. 1981 Jan 22;289(5795):258–262. doi: 10.1038/289258a0. [DOI] [PubMed] [Google Scholar]
  29. Wilhelmsen K. C., Eggleton K., Temin H. M. Nucleic acid sequences of the oncogene v-rel in reticuloendotheliosis virus strain T and its cellular homolog, the proto-oncogene c-rel. J Virol. 1984 Oct;52(1):172–182. doi: 10.1128/jvi.52.1.172-182.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wilhelmsen K. C., Temin H. M. Structure and dimorphism of c-rel (turkey), the cellular homolog to the oncogene of reticuloendotheliosis virus strain T. J Virol. 1984 Feb;49(2):521–529. doi: 10.1128/jvi.49.2.521-529.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Wong T. C., Lai M. M. Avian reticuloendotheliosis virus contains a new class of oncogene of turkey origin. Virology. 1981 May;111(1):289–293. doi: 10.1016/0042-6822(81)90674-7. [DOI] [PubMed] [Google Scholar]
  32. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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