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. 1984 May;50(2):587–597. doi: 10.1128/jvi.50.2.587-597.1984

A 61,000-dalton truncated large T-antigen is uniformly expressed in hamster cells transformed by polyomavirus.

V Rey-Bellet, H Türler
PMCID: PMC255677  PMID: 6323758

Abstract

Various polyomavirus-transformed hamster cell lines derived from tumors or from infected hamster cell cultures synthesized polyoma middle and small tumor (T)-antigens but no full-size large T-antigen. Instead, all cell lines produced the same or similar polyoma T-antigen-related proteins of ca. 61 kilodaltons (kDal). Like large T-antigen synthesized in lytically infected mouse cells, the 61-kDal proteins were phosphoproteins showing electrophoretic and charge heterogeneities. Chromatographic analysis of the methionine-containing tryptic peptides indicated that the 61-kDal proteins were truncated forms of large T-antigen comprising amino acid residues 1 to 485 (+/- 25). Analysis of viral DNA present in hamster chromosomal DNA of three independently isolated cell lines confirmed that synthesis of the 61-kDal proteins was due to a discontinuity in the large T-antigen coding sequence, most likely located between 7 and 8.9 map units on the polyoma DNA map. The three cell lines yielded essentially the same patterns of viral DNA-containing restriction enzyme fragments, suggesting that insertion of viral DNA into the hamster chromosomes took place at closely similar sites.

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Selected References

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

  1. Basilico C., Gattoni S., Zouzias D., Valle G. D. Loss of integrated viral DNA sequences in polyomatransformed cells is associated with an active viral A function. Cell. 1979 Jul;17(3):645–659. doi: 10.1016/0092-8674(79)90272-1. [DOI] [PubMed] [Google Scholar]
  2. Basilico C., Zouzias D., Della-Valle G., Gattoni S., Colantuoni V., Fenton R., Dailey L. Integration and excision of polyoma virus genomes. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):611–620. doi: 10.1101/sqb.1980.044.01.064. [DOI] [PubMed] [Google Scholar]
  3. Bastin M., Bourgaux-Ramoisy D., Bourgaux P. Biological properties of polyoma DNA fragments cloned in plasmid pBR322. J Gen Virol. 1980 Sep;50(1):179–184. doi: 10.1099/0022-1317-50-1-179. [DOI] [PubMed] [Google Scholar]
  4. Benjamin T. L., Schaffhausen B. S., Silver J. E. Polyoma T (tumor) antigen species in abortively and stably transformed cells. J Supramol Struct. 1979;12(1):127–137. doi: 10.1002/jss.400120110. [DOI] [PubMed] [Google Scholar]
  5. Benjamin T. L. The hr-t gene of polyoma virus. Biochim Biophys Acta. 1982 Dec 21;695(2):69–95. doi: 10.1016/0304-419x(82)90018-x. [DOI] [PubMed] [Google Scholar]
  6. Birg F., Dulbecco R., Fried M., Kamen R. State and organization of polyoma virus DNA sequences in transformed rat cell lines. J Virol. 1979 Feb;29(2):633–648. doi: 10.1128/jvi.29.2.633-648.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Botchan M., Topp W., Sambrook J. The arrangement of simian virus 40 sequences in the DNA of transformed cells. Cell. 1976 Oct;9(2):269–287. doi: 10.1016/0092-8674(76)90118-5. [DOI] [PubMed] [Google Scholar]
  8. Chepelinsky A. B., Seif R., Martin R. G. Integration of the simian virus 40 genome into cellular DNA in temperature-sensitive (N) and temperature-insensitive (A) transformants of 3T3 rat and Chinese hamster lung cells. J Virol. 1980 Jul;35(1):184–193. doi: 10.1128/jvi.35.1.184-193.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Chowdhury K., Light S. E., Garon C. F., Ito Y., Israel M. A. A cloned polyoma DNA fragment representing the 5' half of the early gene region is oncogenic. J Virol. 1980 Nov;36(2):566–574. doi: 10.1128/jvi.36.2.566-574.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Chowdhury K., Meltzer M. L., Israel M. A. Stability of polyoma DNA sequences and virus-coded proteins during tumor formation. J Virol. 1982 Mar;41(3):1000–1006. doi: 10.1128/jvi.41.3.1000-1006.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Denhardt D. T. A membrane-filter technique for the detection of complementary DNA. Biochem Biophys Res Commun. 1966 Jun 13;23(5):641–646. doi: 10.1016/0006-291x(66)90447-5. [DOI] [PubMed] [Google Scholar]
  12. Eckhart W., Hutchinson M. A., Hunter T. An activity phosphorylating tyrosine in polyoma T antigen immunoprecipitates. Cell. 1979 Dec;18(4):925–933. doi: 10.1016/0092-8674(79)90205-8. [DOI] [PubMed] [Google Scholar]
  13. Fluck M. M., Benjamin T. L. Comparisons of two early gene functions essential for transformation in polyoma virus and SV-40. Virology. 1979 Jul 15;96(1):205–228. doi: 10.1016/0042-6822(79)90185-5. [DOI] [PubMed] [Google Scholar]
  14. Gross-Bellard M., Oudet P., Chambon P. Isolation of high-molecular-weight DNA from mammalian cells. Eur J Biochem. 1973 Jul 2;36(1):32–38. doi: 10.1111/j.1432-1033.1973.tb02881.x. [DOI] [PubMed] [Google Scholar]
  15. Gélinas C., Bouchard L., Bastin M. Tumorigenic activity of cloned polyoma virus DNA in newborn rats. Experientia. 1981 Oct 15;37(10):1074–1075. doi: 10.1007/BF02085017. [DOI] [PubMed] [Google Scholar]
  16. Hassell J. A., Topp W. C., Rifkin D. B., Moreau P. E. Transformation of rat embryo fibroblasts by cloned polyoma virus DNA fragments containing only part of the early region. Proc Natl Acad Sci U S A. 1980 Jul;77(7):3978–3982. doi: 10.1073/pnas.77.7.3978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Hayday A., Ruley H. E., Fried M. Structural and biological analysis of integrated polyoma virus DNA and its adjacent host sequences cloned from transformed rat cells. J Virol. 1982 Oct;44(1):67–77. doi: 10.1128/jvi.44.1.67-77.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Hutchinson M. A., Hunter T., Eckhart W. Characterization of T antigens in polyoma-infected and transformed cells. Cell. 1978 Sep;15(1):65–77. doi: 10.1016/0092-8674(78)90083-1. [DOI] [PubMed] [Google Scholar]
  19. Israel M. A., Simmons D. T., Hourihan S. L., Rowe W. P., Martin M. A. Interrupting the early region of polyoma virus DNA enhances tumorigenicity. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3713–3716. doi: 10.1073/pnas.76.8.3713. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Israel M. A., Vanderryn D. F., Meltzer M. L., Martin M. A. Characterization of polyoma viral DNA sequences in polyoma-induced hamster tumor cell lines. J Biol Chem. 1980 Apr 25;255(8):3798–3805. [PubMed] [Google Scholar]
  21. Ito Y. Polyoma virus-specific 55K protein isolated from plasma membrane of productively infected cells is virus-coded and important for cell transformation. Virology. 1979 Oct 15;98(1):261–266. doi: 10.1016/0042-6822(79)90545-2. [DOI] [PubMed] [Google Scholar]
  22. Ito Y., Spurr N., Griffin B. E. Middle T antigen as primary inducer of full expression of the phenotype of transformation by polyoma virus. J Virol. 1980 Jul;35(1):219–232. doi: 10.1128/jvi.35.1.219-232.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ito Y., Spurr N. Polyoma virus T antigens expressed in transformed cells: significance of middle T antigen in transformation. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):149–157. doi: 10.1101/sqb.1980.044.01.017. [DOI] [PubMed] [Google Scholar]
  24. Kessler S. W. Rapid isolation of antigens from cells with a staphylococcal protein A-antibody adsorbent: parameters of the interaction of antibody-antigen complexes with protein A. J Immunol. 1975 Dec;115(6):1617–1624. [PubMed] [Google Scholar]
  25. Ketner G., Kelly T. J., Jr Integrated simian virus 40 sequences in transformed cell DNA: analysis using restriction endonucleases. Proc Natl Acad Sci U S A. 1976 Apr;73(4):1102–1106. doi: 10.1073/pnas.73.4.1102. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. 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]
  27. Lania L., Hayday A., Bjursell G., Gandini-Attardi D., Fried M. Organization and expression of integrated polyoma virus DNA sequences in transformed rodent cells. Cold Spring Harb Symp Quant Biol. 1980;44(Pt 1):597–603. doi: 10.1101/sqb.1980.044.01.062. [DOI] [PubMed] [Google Scholar]
  28. Laskey R. A., Mills A. D. Quantitative film detection of 3H and 14C in polyacrylamide gels by fluorography. Eur J Biochem. 1975 Aug 15;56(2):335–341. doi: 10.1111/j.1432-1033.1975.tb02238.x. [DOI] [PubMed] [Google Scholar]
  29. MACPHERSON I., MONTAGNIER L. AGAR SUSPENSION CULTURE FOR THE SELECTIVE ASSAY OF CELLS TRANSFORMED BY POLYOMA VIRUS. Virology. 1964 Jun;23:291–294. doi: 10.1016/0042-6822(64)90301-0. [DOI] [PubMed] [Google Scholar]
  30. Maniatis T., Jeffrey A., Kleid D. G. Nucleotide sequence of the rightward operator of phage lambda. Proc Natl Acad Sci U S A. 1975 Mar;72(3):1184–1188. doi: 10.1073/pnas.72.3.1184. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Marvo S. L., King S. R., Jaskunas S. R. Role of short regions of homology in intermolecular illegitimate recombination events. Proc Natl Acad Sci U S A. 1983 May;80(9):2452–2456. doi: 10.1073/pnas.80.9.2452. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Mes A. M., Hassell J. A. Polyoma viral middle T-antigen is required for transformation. J Virol. 1982 May;42(2):621–629. doi: 10.1128/jvi.42.2.621-629.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Mougneau E., Birg F., Rassoulzadegan M., Cuzin F. Integration sites and sequence arrangement of SV40 DNA in a homogeneous series of transformed rat fibroblast lines. Cell. 1980 Dec;22(3):917–927. doi: 10.1016/0092-8674(80)90569-3. [DOI] [PubMed] [Google Scholar]
  34. Novak U., Dilworth S. M., Griffin B. E. Coding capacity of a 35 percent fragment of the polyoma virus genome is sufficient to initiate and maintain cellular transformation. Proc Natl Acad Sci U S A. 1980 Jun;77(6):3278–3282. doi: 10.1073/pnas.77.6.3278. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. O'Farrell P. H. High resolution two-dimensional electrophoresis of proteins. J Biol Chem. 1975 May 25;250(10):4007–4021. [PMC free article] [PubMed] [Google Scholar]
  36. Pétursson G., Weil R. A study on the mechanism of polyoma-induced activation of the cellular DNA-synthesizing apparatus. Synchronization by FUdR of virus-induced DNA synthesis. Arch Gesamte Virusforsch. 1968;24(1):1–29. doi: 10.1007/BF01242898. [DOI] [PubMed] [Google Scholar]
  37. Rassoulzadegan M., Cowie A., Carr A., Glaichenhaus N., Kamen R., Cuzin F. The roles of individual polyoma virus early proteins in oncogenic transformation. Nature. 1982 Dec 23;300(5894):713–718. doi: 10.1038/300713a0. [DOI] [PubMed] [Google Scholar]
  38. Rigby P. W., Dieckmann M., Rhodes C., Berg P. Labeling deoxyribonucleic acid to high specific activity in vitro by nick translation with DNA polymerase I. J Mol Biol. 1977 Jun 15;113(1):237–251. doi: 10.1016/0022-2836(77)90052-3. [DOI] [PubMed] [Google Scholar]
  39. STOKER M., ABEL P. Conditions affecting transformation by polyoma virus. Cold Spring Harb Symp Quant Biol. 1962;27:375–386. doi: 10.1101/sqb.1962.027.001.035. [DOI] [PubMed] [Google Scholar]
  40. Schaffhausen B. S., Benjamin T. L. Phosphorylation of polyoma T antigens. Cell. 1979 Dec;18(4):935–946. doi: 10.1016/0092-8674(79)90206-x. [DOI] [PubMed] [Google Scholar]
  41. Schaffhausen B. S., Silver J. E., Benjamin T. L. Tumor antigen(s) in cell productively infected by wild-type polyoma virus and mutant NG-18. Proc Natl Acad Sci U S A. 1978 Jan;75(1):79–83. doi: 10.1073/pnas.75.1.79. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Schaffhausen B., Benjamin T. L. Comparison of phosphorylation of two polyoma virus middle T antigens in vivo and in vitro. J Virol. 1981 Oct;40(1):184–196. doi: 10.1128/jvi.40.1.184-196.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Schwyzer M., Weil R., Frank G., Zuber H. Amino acid sequence analysis of fragments generated by partial proteolysis from large simian virus 40 tumor antigen. J Biol Chem. 1980 Jun 25;255(12):5627–5634. [PubMed] [Google Scholar]
  44. Smith A. E., Smith R., Griffin B., Fried M. Protein kinase activity associated with polyoma virus middle T antigen in vitro. Cell. 1979 Dec;18(4):915–924. doi: 10.1016/0092-8674(79)90204-6. [DOI] [PubMed] [Google Scholar]
  45. 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]
  46. Stringer J. R. DNA sequence homology and chromosomal deletion at a site of SV40 DNA integration. Nature. 1982 Mar 25;296(5855):363–366. doi: 10.1038/296363a0. [DOI] [PubMed] [Google Scholar]
  47. Stringer J. R. Integrated simian virus 40 DNA: nucleotide sequences at cell-virus recombinant junctions. J Virol. 1981 May;38(2):671–679. doi: 10.1128/jvi.38.2.671-679.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Templeton D., Eckhart W. Mutation causing premature termination of the polyoma virus medium T antigen blocks cell transformation. J Virol. 1982 Mar;41(3):1014–1024. doi: 10.1128/jvi.41.3.1014-1024.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  49. Treisman R., Novak U., Favaloro J., Kamen R. Transformation of rat cells by an altered polyoma virus genome expressing only the middle-T protein. Nature. 1981 Aug 13;292(5824):595–600. doi: 10.1038/292595a0. [DOI] [PubMed] [Google Scholar]
  50. Türler H. The tumor antigens and the early functions of polyoma virus. Mol Cell Biochem. 1980 Sep 15;32(2):63–93. doi: 10.1007/BF00227801. [DOI] [PubMed] [Google Scholar]
  51. VOGT M., DULBECCO R. Steps in the neoplastic transformation of hamster embryo cells by polyoma virus. Proc Natl Acad Sci U S A. 1963 Feb 15;49:171–179. doi: 10.1073/pnas.49.2.171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Weil R. Viral 'tumor antigens': A novel type of mammalian regulator protein. Biochim Biophys Acta. 1978 Nov 17;516(3):301–388. doi: 10.1016/0304-419x(78)90012-4. [DOI] [PubMed] [Google Scholar]

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