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. 1992 Jan;66(1):53–61. doi: 10.1128/jvi.66.1.53-61.1992

Polyomavirus large and small T antigens cooperate in induction of the S phase in serum-starved 3T3 mouse fibroblasts.

E Ogris 1, I Mudrak 1, E Wintersberger 1
PMCID: PMC238259  PMID: 1309261

Abstract

The induction of an S phase in the host cell is a prerequisite for the lytic replication cycle of polyomavirus. This function was attributed to proteins coded for by the early region of the viral DNA, the T antigens. A consideration of the role of the T antigens in the initiation of a mitogenic response of the host cell has to take into account the recent discovery that virus adsorption is sufficient to induce the synthesis of proteins which are known to appear early after quiescent cells are stimulated by the addition of serum, namely fos, jun, and myc (J. Zullo, C.D. Stiles, and R.L. Garcea, Proc. Natl. Acad. Sci. USA 84:1210-1214, 1987; G. M. Glenn and W. Eckhart, J. Virol. 64:2193-2201, 1990). This induction is followed by an initiation of DNA synthesis. It is therefore important to dissociate the effects of the T antigens on the host cell from those of virus adsorption. To do so, we used dexamethasone-regulated versions of the large and small T antigens of polyomavirus stably integrated into the genome of Swiss 3T3 cells to study their function in S-phase induction. When the production of the large or small T antigen in serum-starved 3T3 mouse fibroblasts was activated, only a small fraction of cells was able to leave G0/G1 despite the synthesis of considerable amounts of the respective T antigen. Activation of both T antigens within the same cell, on the other hand, resulted in S-phase induction in a notable percentage of cells, suggesting that the two proteins cooperate in this activity. Polyomavirus T antigens appear to bypass the pathway of growth regulation involving the activation of c-fos. These results are discussed in relation to other known functions of the two virally coded proteins.

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Selected References

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  1. Asselin C., Bastin M. Sequences from polyomavirus and simian virus 40 large T genes capable of immortalizing primary rat embryo fibroblasts. J Virol. 1985 Dec;56(3):958–968. doi: 10.1128/jvi.56.3.958-968.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Berger H., Wintersberger E. Polyomavirus small T antigen enhances replication of viral genomes in 3T6 mouse fibroblasts. J Virol. 1986 Nov;60(2):768–770. doi: 10.1128/jvi.60.2.768-770.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bikel I., Montano X., Agha M. E., Brown M., McCormack M., Boltax J., Livingston D. M. SV40 small t antigen enhances the transformation activity of limiting concentrations of SV40 large T antigen. Cell. 1987 Jan 30;48(2):321–330. doi: 10.1016/0092-8674(87)90435-1. [DOI] [PubMed] [Google Scholar]
  4. Blochlinger K., Diggelmann H. Hygromycin B phosphotransferase as a selectable marker for DNA transfer experiments with higher eucaryotic cells. Mol Cell Biol. 1984 Dec;4(12):2929–2931. doi: 10.1128/mcb.4.12.2929. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Bockus B. J., Schaffhausen B. Localization of the phosphorylations of polyomavirus large T antigen. J Virol. 1987 Apr;61(4):1155–1163. doi: 10.1128/jvi.61.4.1155-1163.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Bockus B. J., Schaffhausen B. Phosphorylation of polyomavirus large T antigen: effects of viral mutations and cell growth state. J Virol. 1987 Apr;61(4):1147–1154. doi: 10.1128/jvi.61.4.1147-1154.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Boldogh I., AbuBakar S., Albrecht T. Activation of proto-oncogenes: an immediate early event in human cytomegalovirus infection. Science. 1990 Feb 2;247(4942):561–564. doi: 10.1126/science.1689075. [DOI] [PubMed] [Google Scholar]
  8. Boldogh I., AbuBakar S., Deng C. Z., Albrecht T. Transcriptional activation of cellular oncogenes fos, jun, and myc by human cytomegalovirus. J Virol. 1991 Mar;65(3):1568–1571. doi: 10.1128/jvi.65.3.1568-1571.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cheng S. H., Espino P. C., Marshall J., Harvey R., Smith A. E. Stoichiometry of cellular and viral components in the polyomavirus middle-T antigen-tyrosine kinase complex. Mol Cell Biol. 1990 Oct;10(10):5569–5574. doi: 10.1128/mcb.10.10.5569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Cherington V., Morgan B., Spiegelman B. M., Roberts T. M. Recombinant retroviruses that transduce individual polyoma tumor antigens: effects on growth and differentiation. Proc Natl Acad Sci U S A. 1986 Jun;83(12):4307–4311. doi: 10.1073/pnas.83.12.4307. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Cowie A., de Villiers J., Kamen R. Immortalization of rat embryo fibroblasts by mutant polyomavirus large T antigens deficient in DNA binding. Mol Cell Biol. 1986 Dec;6(12):4344–4352. doi: 10.1128/mcb.6.12.4344. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. DeCaprio J. A., Ludlow J. W., Figge J., Shew J. Y., Huang C. M., Lee W. H., Marsilio E., Paucha E., Livingston D. M. SV40 large tumor antigen forms a specific complex with the product of the retinoblastoma susceptibility gene. Cell. 1988 Jul 15;54(2):275–283. doi: 10.1016/0092-8674(88)90559-4. [DOI] [PubMed] [Google Scholar]
  13. Dilworth S. M., Griffin B. E. Monoclonal antibodies against polyoma virus tumor antigens. Proc Natl Acad Sci U S A. 1982 Feb;79(4):1059–1063. doi: 10.1073/pnas.79.4.1059. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Dyson N., Bernards R., Friend S. H., Gooding L. R., Hassell J. A., Major E. O., Pipas J. M., Vandyke T., Harlow E. Large T antigens of many polyomaviruses are able to form complexes with the retinoblastoma protein. J Virol. 1990 Mar;64(3):1353–1356. doi: 10.1128/jvi.64.3.1353-1356.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Eilers M., Picard D., Yamamoto K. R., Bishop J. M. Chimaeras of myc oncoprotein and steroid receptors cause hormone-dependent transformation of cells. Nature. 1989 Jul 6;340(6228):66–68. doi: 10.1038/340066a0. [DOI] [PubMed] [Google Scholar]
  16. Eilers M., Schirm S., Bishop J. M. The MYC protein activates transcription of the alpha-prothymosin gene. EMBO J. 1991 Jan;10(1):133–141. doi: 10.1002/j.1460-2075.1991.tb07929.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  18. Fujimura F. K., Deininger P. L., Friedmann T., Linney E. Mutation near the polyoma DNA replication origin permits productive infection of F9 embryonal carcinoma cells. Cell. 1981 Mar;23(3):809–814. doi: 10.1016/0092-8674(81)90445-1. [DOI] [PubMed] [Google Scholar]
  19. Glenn G. M., Eckhart W. Transcriptional regulation of early-response genes during polyomavirus infection. J Virol. 1990 May;64(5):2193–2201. doi: 10.1128/jvi.64.5.2193-2201.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Griffin B. E., Dilworth S. M. Polyomavirus: an overview of its unique properties. Adv Cancer Res. 1983;39:183–268. doi: 10.1016/s0065-230x(08)61036-2. [DOI] [PubMed] [Google Scholar]
  21. Hamel P. A., Cohen B. L., Sorce L. M., Gallie B. L., Phillips R. A. Hyperphosphorylation of the retinoblastoma gene product is determined by domains outside the simian virus 40 large-T-antigen-binding regions. Mol Cell Biol. 1990 Dec;10(12):6586–6595. doi: 10.1128/mcb.10.12.6586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hassauer M., Scheidtmann K. H., Walter G. Mapping of phosphorylation sites in polyomavirus large T antigen. J Virol. 1986 Jun;58(3):805–816. doi: 10.1128/jvi.58.3.805-816.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Hirt B. Selective extraction of polyoma DNA from infected mouse cell cultures. J Mol Biol. 1967 Jun 14;26(2):365–369. doi: 10.1016/0022-2836(67)90307-5. [DOI] [PubMed] [Google Scholar]
  24. Jat P. S., Sharp P. A. Large T antigens of simian virus 40 and polyomavirus efficiently establish primary fibroblasts. J Virol. 1986 Sep;59(3):746–750. doi: 10.1128/jvi.59.3.746-750.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Kern F. G., Pellegrini S., Cowie A., Basilico C. Regulation of polyomavirus late promoter activity by viral early proteins. J Virol. 1986 Oct;60(1):275–285. doi: 10.1128/jvi.60.1.275-285.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kingston R. E., Cowie A., Morimoto R. I., Gwinn K. A. Binding of polyomavirus large T antigen to the human hsp70 promoter is not required for trans activation. Mol Cell Biol. 1986 Sep;6(9):3180–3190. doi: 10.1128/mcb.6.9.3180. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Labow M. A., Baim S. B., Shenk T., Levine A. J. Conversion of the lac repressor into an allosterically regulated transcriptional activator for mammalian cells. Mol Cell Biol. 1990 Jul;10(7):3343–3356. doi: 10.1128/mcb.10.7.3343. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. 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]
  29. Larose A., Dyson N., Sullivan M., Harlow E., Bastin M. Polyomavirus large T mutants affected in retinoblastoma protein binding are defective in immortalization. J Virol. 1991 May;65(5):2308–2313. doi: 10.1128/jvi.65.5.2308-2313.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Lawson R., Cohen P., Lane D. P. Simian virus 40 large T-antigen-dependent DNA replication is activated by protein phosphatase 2A in vitro. J Virol. 1990 May;64(5):2380–2383. doi: 10.1128/jvi.64.5.2380-2383.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Levine A. J. The p53 protein and its interactions with the oncogene products of the small DNA tumor viruses. Virology. 1990 Aug;177(2):419–426. doi: 10.1016/0042-6822(90)90505-l. [DOI] [PubMed] [Google Scholar]
  32. Linder S., Marshall H. Immortalization of primary cells by DNA tumor viruses. Exp Cell Res. 1990 Nov;191(1):1–7. doi: 10.1016/0014-4827(90)90027-8. [DOI] [PubMed] [Google Scholar]
  33. Loeken M., Bikel I., Livingston D. M., Brady J. trans-activation of RNA polymerase II and III promoters by SV40 small t antigen. Cell. 1988 Dec 23;55(6):1171–1177. doi: 10.1016/0092-8674(88)90261-9. [DOI] [PubMed] [Google Scholar]
  34. Ludlow J. W., Shon J., Pipas J. M., Livingston D. M., DeCaprio J. A. The retinoblastoma susceptibility gene product undergoes cell cycle-dependent dephosphorylation and binding to and release from SV40 large T. Cell. 1990 Feb 9;60(3):387–396. doi: 10.1016/0092-8674(90)90590-b. [DOI] [PubMed] [Google Scholar]
  35. Martens I., Nilsson S. A., Linder S., Magnusson G. Mutational analysis of polyomavirus small-T-antigen functions in productive infection and in transformation. J Virol. 1989 May;63(5):2126–2133. doi: 10.1128/jvi.63.5.2126-2133.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Moran E. A region of SV40 large T antigen can substitute for a transforming domain of the adenovirus E1A products. Nature. 1988 Jul 14;334(6178):168–170. doi: 10.1038/334168a0. [DOI] [PubMed] [Google Scholar]
  37. Morhenn V., Rabinowitz Z., Tomkins G. M. Effects of adrenal glucocorticoids on polyoma virus replication. Proc Natl Acad Sci U S A. 1973 Apr;70(4):1088–1089. doi: 10.1073/pnas.70.4.1088. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Nilsson S. V., Magnusson G. T-antigen expression by polyoma mutants with modified RNA splicing. EMBO J. 1983;2(12):2095–2101. doi: 10.1002/j.1460-2075.1983.tb01708.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Noda T., Satake M., Robins T., Ito Y. Isolation and characterization of NIH 3T3 cells expressing polyomavirus small T antigen. J Virol. 1986 Oct;60(1):105–113. doi: 10.1128/jvi.60.1.105-113.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Pallas D. C., Shahrik L. K., Martin B. L., Jaspers S., Miller T. B., Brautigan D. L., Roberts T. M. Polyoma small and middle T antigens and SV40 small t antigen form stable complexes with protein phosphatase 2A. Cell. 1990 Jan 12;60(1):167–176. doi: 10.1016/0092-8674(90)90726-u. [DOI] [PubMed] [Google Scholar]
  41. Pardee A. B. G1 events and regulation of cell proliferation. Science. 1989 Nov 3;246(4930):603–608. doi: 10.1126/science.2683075. [DOI] [PubMed] [Google Scholar]
  42. Parker B. A., Stark G. R. Regulation of simian virus 40 transcription: sensitive analysis of the RNA species present early in infections by virus or viral DNA. J Virol. 1979 Aug;31(2):360–369. doi: 10.1128/jvi.31.2.360-369.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Raptis L., Lamfrom H., Benjamin T. L. Regulation of cellular phenotype and expression of polyomavirus middle T antigen in rat fibroblasts. Mol Cell Biol. 1985 Sep;5(9):2476–2486. doi: 10.1128/mcb.5.9.2476. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. 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]
  45. Rassoulzadegan M., Naghashfar Z., Cowie A., Carr A., Grisoni M., Kamen R., Cuzin F. Expression of the large T protein of polyoma virus promotes the establishment in culture of "normal" rodent fibroblast cell lines. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4354–4358. doi: 10.1073/pnas.80.14.4354. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. Ray F. A., Peabody D. S., Cooper J. L., Cram L. S., Kraemer P. M. SV40 T antigen alone drives karyotype instability that precedes neoplastic transformation of human diploid fibroblasts. J Cell Biochem. 1990 Jan;42(1):13–31. doi: 10.1002/jcb.240420103. [DOI] [PubMed] [Google Scholar]
  47. Rinehart C. A., Haskill J. S., Morris J. S., Butler T. D., Kaufman D. G. Extended life span of human endometrial stromal cells transfected with cloned origin-defective, temperature-sensitive simian virus 40. J Virol. 1991 Mar;65(3):1458–1465. doi: 10.1128/jvi.65.3.1458-1465.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. 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]
  49. St-Onge L., Bouchard L., Laurent S., Bastin M. Intrachromosomal recombination mediated by papovavirus large T antigens. J Virol. 1990 Jun;64(6):2958–2966. doi: 10.1128/jvi.64.6.2958-2966.1990. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Stewart N., Bacchetti S. Expression of SV40 large T antigen, but not small t antigen, is required for the induction of chromosomal aberrations in transformed human cells. Virology. 1991 Jan;180(1):49–57. doi: 10.1016/0042-6822(91)90008-y. [DOI] [PubMed] [Google Scholar]
  51. Strauss M., Griffin B. E. Cellular immortalization--an essential step or merely a risk factor in DNA virus-induced transformation? Cancer Cells. 1990 Nov;2(11):360–365. [PubMed] [Google Scholar]
  52. Strauss M., Hering S., Lubbe L., Griffin B. E. Immortalization and transformation of human fibroblasts by regulated expression of polyoma virus T antigens. Oncogene. 1990 Aug;5(8):1223–1229. [PubMed] [Google Scholar]
  53. Templeton D., Simon S., Eckhart W. Truncated forms of the polyomavirus middle T antigen can substitute for the small T antigen in lytic infection. J Virol. 1986 Jan;57(1):367–370. doi: 10.1128/jvi.57.1.367-370.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  54. Türler H., Salomon C. Small and middle T antigens contribute to lytic and abortive polyomavirus infection. J Virol. 1985 Feb;53(2):579–586. doi: 10.1128/jvi.53.2.579-586.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Virshup D. M., Kauffman M. G., Kelly T. J. Activation of SV40 DNA replication in vitro by cellular protein phosphatase 2A. EMBO J. 1989 Dec 1;8(12):3891–3898. doi: 10.1002/j.1460-2075.1989.tb08568.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Walter G., Ruediger R., Slaughter C., Mumby M. Association of protein phosphatase 2A with polyoma virus medium tumor antigen. Proc Natl Acad Sci U S A. 1990 Apr;87(7):2521–2525. doi: 10.1073/pnas.87.7.2521. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Wigler M., Pellicer A., Silverstein S., Axel R. Biochemical transfer of single-copy eucaryotic genes using total cellular DNA as donor. Cell. 1978 Jul;14(3):725–731. doi: 10.1016/0092-8674(78)90254-4. [DOI] [PubMed] [Google Scholar]
  58. Zhu Z. Y., Veldman G. M., Cowie A., Carr A., Schaffhausen B., Kamen R. Construction and functional characterization of polyomavirus genomes that separately encode the three early proteins. J Virol. 1984 Jul;51(1):170–180. doi: 10.1128/jvi.51.1.170-180.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Zullo J., Stiles C. D., Garcea R. L. Regulation of c-myc and c-fos mRNA levels by polyomavirus: distinct roles for the capsid protein VP1 and the viral early proteins. Proc Natl Acad Sci U S A. 1987 Mar;84(5):1210–1214. doi: 10.1073/pnas.84.5.1210. [DOI] [PMC free article] [PubMed] [Google Scholar]

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