Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1986 Sep;6(9):3180–3190. doi: 10.1128/mcb.6.9.3180

Binding of polyomavirus large T antigen to the human hsp70 promoter is not required for trans activation.

R E Kingston, A Cowie, R I Morimoto, K A Gwinn
PMCID: PMC367054  PMID: 3023966

Abstract

Polyomavirus large T antigen binds to two sites located between positions -110 and -170 of a human heat shock protein 70 (hsp70) promoter. Methylation interference studies show that binding for each site is determined by two GPuGGC pentanucleotide sequences. The specificity of this binding interaction is similar to that observed for large T binding to the viral genome. The existence of sequences that bind a viral protein in a cellular promoter raises the possibility that these sequences play a role in gene expression in an uninfected cell. We show that hsp70 large T antigen binding site 1 is capable of functioning as an upstream promoter element in cells that do not contain any viral T antigen. Genetic analysis of this effect suggests that a cellular factor exists that has a binding specificity that overlaps but is not identical to that of polyomavirus large T antigen. To determine whether binding of polyomavirus large T antigen can regulate expression of the intact human hsp70 promoter, we have introduced the promoter into mouse cells with plasmids that express the polyomavirus early proteins. These proteins stimulate the level of correctly initiated hsp70 transcripts, but surprisingly the degree of stimulation remains unchanged for promoter constructs in which the large T antigen binding sites have been deleted. These observations suggest that trans activation of the hsp70 promoter by the polyomavirus early proteins occurs through protein-protein interactions and not through sequence-specific DNA binding.

Full text

PDF
3184

Images in this article

3182Image
on p.3182
3183Image
on p.3183
3183Image
on p.3183
3186Image
on p.3186
3187Image
on p.3187

Selected References

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

  1. Alwine J. C. Transient gene expression control: effects of transfected DNA stability and trans-activation by viral early proteins. Mol Cell Biol. 1985 May;5(5):1034–1042. doi: 10.1128/mcb.5.5.1034. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Bardwell J. C., Craig E. A. Major heat shock gene of Drosophila and the Escherichia coli heat-inducible dnaK gene are homologous. Proc Natl Acad Sci U S A. 1984 Feb;81(3):848–852. doi: 10.1073/pnas.81.3.848. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bensaude O., Babinet C., Morange M., Jacob F. Heat shock proteins, first major products of zygotic gene activity in mouse embryo. Nature. 1983 Sep 22;305(5932):331–333. doi: 10.1038/305331a0. [DOI] [PubMed] [Google Scholar]
  4. Berk A. J., Lee F., Harrison T., Williams J., Sharp P. A. Pre-early adenovirus 5 gene product regulates synthesis of early viral messenger RNAs. Cell. 1979 Aug;17(4):935–944. doi: 10.1016/0092-8674(79)90333-7. [DOI] [PubMed] [Google Scholar]
  5. Berk A. J., Sharp P. A. Sizing and mapping of early adenovirus mRNAs by gel electrophoresis of S1 endonuclease-digested hybrids. Cell. 1977 Nov;12(3):721–732. doi: 10.1016/0092-8674(77)90272-0. [DOI] [PubMed] [Google Scholar]
  6. Brady J., Bolen J. B., Radonovich M., Salzman N., Khoury G. Stimulation of simian virus 40 late gene expression by simian virus 40 tumor antigen. Proc Natl Acad Sci U S A. 1984 Apr;81(7):2040–2044. doi: 10.1073/pnas.81.7.2040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Brady J., Khoury G. trans Activation of the simian virus 40 late transcription unit by T-antigen. Mol Cell Biol. 1985 Jun;5(6):1391–1399. doi: 10.1128/mcb.5.6.1391. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Brady J., Loeken M. R., Khoury G. Interaction between two transcriptional control sequences required for tumor-antigen-mediated simian virus 40 late gene expression. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7299–7303. doi: 10.1073/pnas.82.21.7299. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Cepko C. L., Roberts B. E., Mulligan R. C. Construction and applications of a highly transmissible murine retrovirus shuttle vector. Cell. 1984 Jul;37(3):1053–1062. doi: 10.1016/0092-8674(84)90440-9. [DOI] [PubMed] [Google Scholar]
  10. Chen E. Y., Seeburg P. H. Supercoil sequencing: a fast and simple method for sequencing plasmid DNA. DNA. 1985 Apr;4(2):165–170. doi: 10.1089/dna.1985.4.165. [DOI] [PubMed] [Google Scholar]
  11. Cowie A., Kamen R. Guanine nucleotide contacts within viral DNA sequences bound by polyomavirus large T antigen. J Virol. 1986 Feb;57(2):505–514. doi: 10.1128/jvi.57.2.505-514.1986. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Cowie A., Kamen R. Multiple binding sites for polyomavirus large T antigen within regulatory sequences of polyomavirus DNA. J Virol. 1984 Dec;52(3):750–760. doi: 10.1128/jvi.52.3.750-760.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Craig E. A., Jacobsen K. Mutations in cognate genes of Saccharomyces cerevisiae hsp70 result in reduced growth rates at low temperatures. Mol Cell Biol. 1985 Dec;5(12):3517–3524. doi: 10.1128/mcb.5.12.3517. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Craig E. A., Jacobsen K. Mutations of the heat inducible 70 kilodalton genes of yeast confer temperature sensitive growth. Cell. 1984 Oct;38(3):841–849. doi: 10.1016/0092-8674(84)90279-4. [DOI] [PubMed] [Google Scholar]
  15. DeLucia A. L., Lewton B. A., Tjian R., Tegtmeyer P. Topography of simian virus 40 A protein-DNA complexes: arrangement of pentanucleotide interaction sites at the origin of replication. J Virol. 1983 Apr;46(1):143–150. doi: 10.1128/jvi.46.1.143-150.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Dilworth S. M., Cowie A., Kamen R. I., Griffin B. E. DNA binding activity of polyoma virus large tumor antigen. Proc Natl Acad Sci U S A. 1984 Apr;81(7):1941–1945. doi: 10.1073/pnas.81.7.1941. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Efstratiadis A., Posakony J. W., Maniatis T., Lawn R. M., O'Connell C., Spritz R. A., DeRiel J. K., Forget B. G., Weissman S. M., Slightom J. L. The structure and evolution of the human beta-globin gene family. Cell. 1980 Oct;21(3):653–668. doi: 10.1016/0092-8674(80)90429-8. [DOI] [PubMed] [Google Scholar]
  18. Galas D. J., Schmitz A. DNAse footprinting: a simple method for the detection of protein-DNA binding specificity. Nucleic Acids Res. 1978 Sep;5(9):3157–3170. doi: 10.1093/nar/5.9.3157. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Gaudray P., Tyndall C., Kamen R., Cuzin F. The high affinity binding site on polyoma virus DNA for the viral large-T protein. Nucleic Acids Res. 1981 Nov 11;9(21):5697–5710. doi: 10.1093/nar/9.21.5697. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Gaynor R. B., Hillman D., Berk A. J. Adenovirus early region 1A protein activates transcription of a nonviral gene introduced into mammalian cells by infection or transfection. Proc Natl Acad Sci U S A. 1984 Feb;81(4):1193–1197. doi: 10.1073/pnas.81.4.1193. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Gorman C. M., Moffat L. F., Howard B. H. Recombinant genomes which express chloramphenicol acetyltransferase in mammalian cells. Mol Cell Biol. 1982 Sep;2(9):1044–1051. doi: 10.1128/mcb.2.9.1044. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. 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]
  23. Graves B. J., Johnson P. F., McKnight S. L. Homologous recognition of a promoter domain common to the MSV LTR and the HSV tk gene. Cell. 1986 Feb 28;44(4):565–576. doi: 10.1016/0092-8674(86)90266-7. [DOI] [PubMed] [Google Scholar]
  24. Green M. R., Treisman R., Maniatis T. Transcriptional activation of cloned human beta-globin genes by viral immediate-early gene products. Cell. 1983 Nov;35(1):137–148. doi: 10.1016/0092-8674(83)90216-7. [DOI] [PubMed] [Google Scholar]
  25. Houweling A., van den Elsen P. J., van der Eb A. J. Partial transformation of primary rat cells by the leftmost 4.5% fragment of adenovirus 5 DNA. Virology. 1980 Sep;105(2):537–550. doi: 10.1016/0042-6822(80)90054-9. [DOI] [PubMed] [Google Scholar]
  26. Hunt C., Morimoto R. I. Conserved features of eukaryotic hsp70 genes revealed by comparison with the nucleotide sequence of human hsp70. Proc Natl Acad Sci U S A. 1985 Oct;82(19):6455–6459. doi: 10.1073/pnas.82.19.6455. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Imperiale M. J., Kao H. T., Feldman L. T., Nevins J. R., Strickland S. Common control of the heat shock gene and early adenovirus genes: evidence for a cellular E1A-like activity. Mol Cell Biol. 1984 May;4(5):867–874. doi: 10.1128/mcb.4.5.867. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Jat P. S., Cepko C. L., Mulligan R. C., Sharp P. A. Recombinant retroviruses encoding simian virus 40 large T antigen and polyomavirus large and middle T antigens. Mol Cell Biol. 1986 Apr;6(4):1204–1217. doi: 10.1128/mcb.6.4.1204. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Jones K. A., Tjian R. Essential contact residues within SV40 large T antigen binding sites I and II identified by alkylation-interference. Cell. 1984 Jan;36(1):155–162. doi: 10.1016/0092-8674(84)90084-9. [DOI] [PubMed] [Google Scholar]
  30. Jones N., Shenk T. An adenovirus type 5 early gene function regulates expression of other early viral genes. Proc Natl Acad Sci U S A. 1979 Aug;76(8):3665–3669. doi: 10.1073/pnas.76.8.3665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Kao H. T., Nevins J. R. Transcriptional activation and subsequent control of the human heat shock gene during adenovirus infection. Mol Cell Biol. 1983 Nov;3(11):2058–2065. doi: 10.1128/mcb.3.11.2058. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Keller J. M., Alwine J. C. Activation of the SV40 late promoter: direct effects of T antigen in the absence of viral DNA replication. Cell. 1984 Feb;36(2):381–389. doi: 10.1016/0092-8674(84)90231-9. [DOI] [PubMed] [Google Scholar]
  33. Keller J. M., Alwine J. C. Analysis of an activatable promoter: sequences in the simian virus 40 late promoter required for T-antigen-mediated trans activation. Mol Cell Biol. 1985 Aug;5(8):1859–1869. doi: 10.1128/mcb.5.8.1859. [DOI] [PMC free article] [PubMed] [Google Scholar]
  34. Khandjian E. W., Türler H. Simian virus 40 and polyoma virus induce synthesis of heat shock proteins in permissive cells. Mol Cell Biol. 1983 Jan;3(1):1–8. doi: 10.1128/mcb.3.1.1. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Kingston R. E., Baldwin A. S., Jr, Sharp P. A. Regulation of heat shock protein 70 gene expression by c-myc. Nature. 1984 Nov 15;312(5991):280–282. doi: 10.1038/312280a0. [DOI] [PubMed] [Google Scholar]
  36. Kingston R. E., Baldwin A. S., Sharp P. A. Transcription control by oncogenes. Cell. 1985 May;41(1):3–5. doi: 10.1016/0092-8674(85)90049-2. [DOI] [PubMed] [Google Scholar]
  37. Kingston R. E., Kaufman R. J., Sharp P. A. Regulation of transcription of the adenovirus EII promoter by EIa gene products: absence of sequence specificity. Mol Cell Biol. 1984 Oct;4(10):1970–1977. doi: 10.1128/mcb.4.10.1970. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Lanford R. E., Wong C., Butel J. S. Differential ability of a T-antigen transport-defective mutant of simian virus 40 to transform primary and established rodent cells. Mol Cell Biol. 1985 May;5(5):1043–1050. doi: 10.1128/mcb.5.5.1043. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Matter J. M., Tiercy J. M., Weil R. Sequential stimulation of cellular RNA synthesis in polyoma-infected mouse kidney cell cultures. Nucleic Acids Res. 1983 Oct 11;11(19):6611–6629. doi: 10.1093/nar/11.19.6611. [DOI] [PMC free article] [PubMed] [Google Scholar]
  40. Maxam A. M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. Methods Enzymol. 1980;65(1):499–560. doi: 10.1016/s0076-6879(80)65059-9. [DOI] [PubMed] [Google Scholar]
  41. Murthy S. C., Bhat G. P., Thimmappaya B. Adenovirus EIIA early promoter: transcriptional control elements and induction by the viral pre-early EIA gene, which appears to be sequence independent. Proc Natl Acad Sci U S A. 1985 Apr;82(8):2230–2234. doi: 10.1073/pnas.82.8.2230. [DOI] [PMC free article] [PubMed] [Google Scholar]
  42. Myers R. M., Tilly K., Maniatis T. Fine structure genetic analysis of a beta-globin promoter. Science. 1986 May 2;232(4750):613–618. doi: 10.1126/science.3457470. [DOI] [PubMed] [Google Scholar]
  43. Nevins J. R. Induction of the synthesis of a 70,000 dalton mammalian heat shock protein by the adenovirus E1A gene product. Cell. 1982 Jul;29(3):913–919. doi: 10.1016/0092-8674(82)90453-6. [DOI] [PubMed] [Google Scholar]
  44. Petit C. A., Gardes M., Feunteun J. Immortalization of rodent embryo fibroblasts by SV40 is maintained by the A gene. Virology. 1983 May;127(1):74–82. doi: 10.1016/0042-6822(83)90372-0. [DOI] [PubMed] [Google Scholar]
  45. Pomerantz B. J., Hassell J. A. Polyomavirus and simian virus 40 large T antigens bind to common DNA sequences. J Virol. 1984 Mar;49(3):925–937. doi: 10.1128/jvi.49.3.925-937.1984. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. 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]
  47. 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]
  48. Ryder K., Vakalopoulou E., Mertz R., Mastrangelo I., Hough P., Tegtmeyer P., Fanning E. Seventeen base pairs of region I encode a novel tripartite binding signal for SV40 T antigen. Cell. 1985 Sep;42(2):539–548. doi: 10.1016/0092-8674(85)90111-4. [DOI] [PubMed] [Google Scholar]
  49. Sanger F., Nicklen S., Coulson A. R. DNA sequencing with chain-terminating inhibitors. Proc Natl Acad Sci U S A. 1977 Dec;74(12):5463–5467. doi: 10.1073/pnas.74.12.5463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  50. Scott M. R., Westphal K. H., Rigby P. W. Activation of mouse genes in transformed cells. Cell. 1983 Sep;34(2):557–567. doi: 10.1016/0092-8674(83)90388-4. [DOI] [PubMed] [Google Scholar]
  51. Selden R. F., Howie K. B., Rowe M. E., Goodman H. M., Moore D. D. Human growth hormone as a reporter gene in regulation studies employing transient gene expression. Mol Cell Biol. 1986 Sep;6(9):3173–3179. doi: 10.1128/mcb.6.9.3173. [DOI] [PMC free article] [PubMed] [Google Scholar]
  52. Tegtmeyer P., Andersen B., Shaw S. B., Wilson V. G. Alternative interactions of the SV40 A protein with DNA. Virology. 1981 Nov;115(1):75–87. doi: 10.1016/0042-6822(81)90090-8. [DOI] [PubMed] [Google Scholar]
  53. Tjian R. Protein-DNA interactions at the origin of simian virus 40 DNA replication. Cold Spring Harb Symp Quant Biol. 1979;43(Pt 2):655–661. doi: 10.1101/sqb.1979.043.01.073. [DOI] [PubMed] [Google Scholar]
  54. Ullrich A., Shine J., Chirgwin J., Pictet R., Tischer E., Rutter W. J., Goodman H. M. Rat insulin genes: construction of plasmids containing the coding sequences. Science. 1977 Jun 17;196(4296):1313–1319. doi: 10.1126/science.325648. [DOI] [PubMed] [Google Scholar]
  55. Wright P. J., DeLucia A. L., Tegtmeyer P. Sequence-specific binding of simian virus 40 A protein to nonorigin and cellular DNA. Mol Cell Biol. 1984 Dec;4(12):2631–2638. doi: 10.1128/mcb.4.12.2631. [DOI] [PMC free article] [PubMed] [Google Scholar]
  56. Wu B. J., Hurst H. C., Jones N. C., Morimoto R. I. The E1A 13S product of adenovirus 5 activates transcription of the cellular human HSP70 gene. Mol Cell Biol. 1986 Aug;6(8):2994–2999. doi: 10.1128/mcb.6.8.2994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Wu B. J., Kingston R. E., Morimoto R. I. Human HSP70 promoter contains at least two distinct regulatory domains. Proc Natl Acad Sci U S A. 1986 Feb;83(3):629–633. doi: 10.1073/pnas.83.3.629. [DOI] [PMC free article] [PubMed] [Google Scholar]
  58. Wu B. J., Morimoto R. I. Transcription of the human hsp70 gene is induced by serum stimulation. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6070–6074. doi: 10.1073/pnas.82.18.6070. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Wu B., Hunt C., Morimoto R. Structure and expression of the human gene encoding major heat shock protein HSP70. Mol Cell Biol. 1985 Feb;5(2):330–341. doi: 10.1128/mcb.5.2.330. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. Zajchowski D. A., Boeuf H., Kédinger C. The adenovirus-2 early EIIa transcription unit possesses two overlapping promoters with different sequence requirements for EIa-dependent stimulation. EMBO J. 1985 May;4(5):1293–1300. doi: 10.1002/j.1460-2075.1985.tb03775.x. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Molecular and Cellular Biology are provided here courtesy of Taylor & Francis

RESOURCES