Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1993 Sep;13(9):5427–5438. doi: 10.1128/mcb.13.9.5427

The DNA-binding activity of the human heat shock transcription factor is regulated in vivo by hsp70.

D D Mosser 1, J Duchaine 1, B Massie 1
PMCID: PMC360250  PMID: 8355691

Abstract

The human heat shock transcription factor (HSF) is maintained in an inactive non-DNA-binding form under nonstress conditions and acquires the ability to bind specifically to the heat shock promoter element in response to elevated temperatures or other conditions that disrupt protein structure. Here we show that constitutive overexpression of the major inducible heat shock protein, hsp70, in transfected human cells reduces the extent of HSF activation after a heat stress. HSF activation was inhibited more strongly in clones that express higher levels of hsp70. These results demonstrate that HSF activity is negatively regulated in vivo by hsp70 and suggest that the cell might sense elevated temperature as a decreased availability of hsp70. HSF activation in response to treatment with sodium arsenite or the proline analog azetidine was also depressed in hsp70-expressing cells relative to that in the nontransfected control cells. As well, the level of activated HSF decreased more rapidly in the hsp70-expressing clones when the cells were heat shocked and returned to 37 degrees C. These results suggest that hsp70 could play an active role in the conversion of HSF back to a conformation that does not bind the heat shock promoter element during the attenuation of the heat shock response.

Full text

PDF
5427

Images in this article

5429Image
on p.5429
5431Image
on p.5431
5432Image
on p.5432
5432Image
on p.5432
5433Image
on p.5433
5433Image
on p.5433
5434Image
on p.5434

Selected References

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

  1. Abravaya K., Myers M. P., Murphy S. P., Morimoto R. I. The human heat shock protein hsp70 interacts with HSF, the transcription factor that regulates heat shock gene expression. Genes Dev. 1992 Jul;6(7):1153–1164. doi: 10.1101/gad.6.7.1153. [DOI] [PubMed] [Google Scholar]
  2. Abravaya K., Phillips B., Morimoto R. I. Attenuation of the heat shock response in HeLa cells is mediated by the release of bound heat shock transcription factor and is modulated by changes in growth and in heat shock temperatures. Genes Dev. 1991 Nov;5(11):2117–2127. doi: 10.1101/gad.5.11.2117. [DOI] [PubMed] [Google Scholar]
  3. Amici C., Sistonen L., Santoro M. G., Morimoto R. I. Antiproliferative prostaglandins activate heat shock transcription factor. Proc Natl Acad Sci U S A. 1992 Jul 15;89(14):6227–6231. doi: 10.1073/pnas.89.14.6227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Baler R., Welch W. J., Voellmy R. Heat shock gene regulation by nascent polypeptides and denatured proteins: hsp70 as a potential autoregulatory factor. J Cell Biol. 1992 Jun;117(6):1151–1159. doi: 10.1083/jcb.117.6.1151. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Beckmann R. P., Lovett M., Welch W. J. Examining the function and regulation of hsp 70 in cells subjected to metabolic stress. J Cell Biol. 1992 Jun;117(6):1137–1150. doi: 10.1083/jcb.117.6.1137. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Beckmann R. P., Mizzen L. E., Welch W. J. Interaction of Hsp 70 with newly synthesized proteins: implications for protein folding and assembly. Science. 1990 May 18;248(4957):850–854. doi: 10.1126/science.2188360. [DOI] [PubMed] [Google Scholar]
  7. Boorstein W. R., Craig E. A. Transcriptional regulation of SSA3, an HSP70 gene from Saccharomyces cerevisiae. Mol Cell Biol. 1990 Jun;10(6):3262–3267. doi: 10.1128/mcb.10.6.3262. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Clos J., Westwood J. T., Becker P. B., Wilson S., Lambert K., Wu C. Molecular cloning and expression of a hexameric Drosophila heat shock factor subject to negative regulation. Cell. 1990 Nov 30;63(5):1085–1097. doi: 10.1016/0092-8674(90)90511-c. [DOI] [PubMed] [Google Scholar]
  9. Craig E. A., Gross C. A. Is hsp70 the cellular thermometer? Trends Biochem Sci. 1991 Apr;16(4):135–140. doi: 10.1016/0968-0004(91)90055-z. [DOI] [PubMed] [Google Scholar]
  10. 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]
  11. DiDomenico B. J., Bugaisky G. E., Lindquist S. The heat shock response is self-regulated at both the transcriptional and posttranscriptional levels. Cell. 1982 Dec;31(3 Pt 2):593–603. doi: 10.1016/0092-8674(82)90315-4. [DOI] [PubMed] [Google Scholar]
  12. Ey P. L., Ashman L. K. The use of alkaline phosphatase-conjugated anti-immunoglobulin with immunoblots for determining the specificity of monoclonal antibodies to protein mixtures. Methods Enzymol. 1986;121:497–509. doi: 10.1016/0076-6879(86)21050-2. [DOI] [PubMed] [Google Scholar]
  13. Feder J. H., Rossi J. M., Solomon J., Solomon N., Lindquist S. The consequences of expressing hsp70 in Drosophila cells at normal temperatures. Genes Dev. 1992 Aug;6(8):1402–1413. doi: 10.1101/gad.6.8.1402. [DOI] [PubMed] [Google Scholar]
  14. Gallo G. J., Prentice H., Kingston R. E. Heat shock factor is required for growth at normal temperatures in the fission yeast Schizosaccharomyces pombe. Mol Cell Biol. 1993 Feb;13(2):749–761. doi: 10.1128/mcb.13.2.749. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Gamer J., Bujard H., Bukau B. Physical interaction between heat shock proteins DnaK, DnaJ, and GrpE and the bacterial heat shock transcription factor sigma 32. Cell. 1992 May 29;69(5):833–842. doi: 10.1016/0092-8674(92)90294-m. [DOI] [PubMed] [Google Scholar]
  16. Gething M. J., Sambrook J. Protein folding in the cell. Nature. 1992 Jan 2;355(6355):33–45. doi: 10.1038/355033a0. [DOI] [PubMed] [Google Scholar]
  17. Grossman A. D., Straus D. B., Walter W. A., Gross C. A. Sigma 32 synthesis can regulate the synthesis of heat shock proteins in Escherichia coli. Genes Dev. 1987 Apr;1(2):179–184. doi: 10.1101/gad.1.2.179. [DOI] [PubMed] [Google Scholar]
  18. Gunning P., Leavitt J., Muscat G., Ng S. Y., Kedes L. A human beta-actin expression vector system directs high-level accumulation of antisense transcripts. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4831–4835. doi: 10.1073/pnas.84.14.4831. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Hartl F. U., Martin J., Neupert W. Protein folding in the cell: the role of molecular chaperones Hsp70 and Hsp60. Annu Rev Biophys Biomol Struct. 1992;21:293–322. doi: 10.1146/annurev.bb.21.060192.001453. [DOI] [PubMed] [Google Scholar]
  20. 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]
  21. Jakobsen B. K., Pelham H. R. A conserved heptapeptide restrains the activity of the yeast heat shock transcription factor. EMBO J. 1991 Feb;10(2):369–375. doi: 10.1002/j.1460-2075.1991.tb07958.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Jakobsen B. K., Pelham H. R. Constitutive binding of yeast heat shock factor to DNA in vivo. Mol Cell Biol. 1988 Nov;8(11):5040–5042. doi: 10.1128/mcb.8.11.5040. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Jättelä M., Wissing D., Bauer P. A., Li G. C. Major heat shock protein hsp70 protects tumor cells from tumor necrosis factor cytotoxicity. EMBO J. 1992 Oct;11(10):3507–3512. doi: 10.1002/j.1460-2075.1992.tb05433.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Kingston R. E., Schuetz T. J., Larin Z. Heat-inducible human factor that binds to a human hsp70 promoter. Mol Cell Biol. 1987 Apr;7(4):1530–1534. doi: 10.1128/mcb.7.4.1530. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Larson J. S., Schuetz T. J., Kingston R. E. Activation in vitro of sequence-specific DNA binding by a human regulatory factor. Nature. 1988 Sep 22;335(6188):372–375. doi: 10.1038/335372a0. [DOI] [PubMed] [Google Scholar]
  26. Li G. C., Li L. G., Liu Y. K., Mak J. Y., Chen L. L., Lee W. M. Thermal response of rat fibroblasts stably transfected with the human 70-kDa heat shock protein-encoding gene. Proc Natl Acad Sci U S A. 1991 Mar 1;88(5):1681–1685. doi: 10.1073/pnas.88.5.1681. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Liberek K., Galitski T. P., Zylicz M., Georgopoulos C. The DnaK chaperone modulates the heat shock response of Escherichia coli by binding to the sigma 32 transcription factor. Proc Natl Acad Sci U S A. 1992 Apr 15;89(8):3516–3520. doi: 10.1073/pnas.89.8.3516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Liu R. Y., Li X., Li L., Li G. C. Expression of human hsp70 in rat fibroblasts enhances cell survival and facilitates recovery from translational and transcriptional inhibition following heat shock. Cancer Res. 1992 Jul 1;52(13):3667–3673. [PubMed] [Google Scholar]
  29. Mizzen L. A., Welch W. J. Characterization of the thermotolerant cell. I. Effects on protein synthesis activity and the regulation of heat-shock protein 70 expression. J Cell Biol. 1988 Apr;106(4):1105–1116. doi: 10.1083/jcb.106.4.1105. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Morimoto R. I., Sarge K. D., Abravaya K. Transcriptional regulation of heat shock genes. A paradigm for inducible genomic responses. J Biol Chem. 1992 Nov 5;267(31):21987–21990. [PubMed] [Google Scholar]
  31. Mosser D. D., Kotzbauer P. T., Sarge K. D., Morimoto R. I. In vitro activation of heat shock transcription factor DNA-binding by calcium and biochemical conditions that affect protein conformation. Proc Natl Acad Sci U S A. 1990 May;87(10):3748–3752. doi: 10.1073/pnas.87.10.3748. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Mosser D. D., Theodorakis N. G., Morimoto R. I. Coordinate changes in heat shock element-binding activity and HSP70 gene transcription rates in human cells. Mol Cell Biol. 1988 Nov;8(11):4736–4744. doi: 10.1128/mcb.8.11.4736. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Nadeau K., Das A., Walsh C. T. Hsp90 chaperonins possess ATPase activity and bind heat shock transcription factors and peptidyl prolyl isomerases. J Biol Chem. 1993 Jan 15;268(2):1479–1487. [PubMed] [Google Scholar]
  34. Nieto-Sotelo J., Wiederrecht G., Okuda A., Parker C. S. The yeast heat shock transcription factor contains a transcriptional activation domain whose activity is repressed under nonshock conditions. Cell. 1990 Aug 24;62(4):807–817. doi: 10.1016/0092-8674(90)90124-w. [DOI] [PubMed] [Google Scholar]
  35. Palleros D. R., Welch W. J., Fink A. L. Interaction of hsp70 with unfolded proteins: effects of temperature and nucleotides on the kinetics of binding. Proc Natl Acad Sci U S A. 1991 Jul 1;88(13):5719–5723. doi: 10.1073/pnas.88.13.5719. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Price B. D., Calderwood S. K. Heat-induced transcription from RNA polymerases II and III and HSF binding activity are co-ordinately regulated by the products of the heat shock genes. J Cell Physiol. 1992 Nov;153(2):392–401. doi: 10.1002/jcp.1041530219. [DOI] [PubMed] [Google Scholar]
  37. Rabindran S. K., Giorgi G., Clos J., Wu C. Molecular cloning and expression of a human heat shock factor, HSF1. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):6906–6910. doi: 10.1073/pnas.88.16.6906. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. Rabindran S. K., Haroun R. I., Clos J., Wisniewski J., Wu C. Regulation of heat shock factor trimer formation: role of a conserved leucine zipper. Science. 1993 Jan 8;259(5092):230–234. doi: 10.1126/science.8421783. [DOI] [PubMed] [Google Scholar]
  39. Ravid Z., Goldblum N., Zaizov R., Schlesinger M., Kertes T., Minowada J., Verbi W., Greaves M. Establishment and characterization of a new leukaemic T-cell line (Peer) with an unusual phenotype. Int J Cancer. 1980 Jun 15;25(6):705–710. doi: 10.1002/ijc.2910250604. [DOI] [PubMed] [Google Scholar]
  40. Sarge K. D., Murphy S. P., Morimoto R. I. Activation of heat shock gene transcription by heat shock factor 1 involves oligomerization, acquisition of DNA-binding activity, and nuclear localization and can occur in the absence of stress. Mol Cell Biol. 1993 Mar;13(3):1392–1407. doi: 10.1128/mcb.13.3.1392. [DOI] [PMC free article] [PubMed] [Google Scholar]
  41. Sarge K. D., Zimarino V., Holm K., Wu C., Morimoto R. I. Cloning and characterization of two mouse heat shock factors with distinct inducible and constitutive DNA-binding ability. Genes Dev. 1991 Oct;5(10):1902–1911. doi: 10.1101/gad.5.10.1902. [DOI] [PubMed] [Google Scholar]
  42. Schuetz T. J., Gallo G. J., Sheldon L., Tempst P., Kingston R. E. Isolation of a cDNA for HSF2: evidence for two heat shock factor genes in humans. Proc Natl Acad Sci U S A. 1991 Aug 15;88(16):6911–6915. doi: 10.1073/pnas.88.16.6911. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. Sorger P. K. Heat shock factor and the heat shock response. Cell. 1991 May 3;65(3):363–366. doi: 10.1016/0092-8674(91)90452-5. [DOI] [PubMed] [Google Scholar]
  44. Sorger P. K., Lewis M. J., Pelham H. R. Heat shock factor is regulated differently in yeast and HeLa cells. Nature. 1987 Sep 3;329(6134):81–84. doi: 10.1038/329081a0. [DOI] [PubMed] [Google Scholar]
  45. Sorger P. K., Nelson H. C. Trimerization of a yeast transcriptional activator via a coiled-coil motif. Cell. 1989 Dec 1;59(5):807–813. doi: 10.1016/0092-8674(89)90604-1. [DOI] [PubMed] [Google Scholar]
  46. Sorger P. K., Pelham H. R. Yeast heat shock factor is an essential DNA-binding protein that exhibits temperature-dependent phosphorylation. Cell. 1988 Sep 9;54(6):855–864. doi: 10.1016/s0092-8674(88)91219-6. [DOI] [PubMed] [Google Scholar]
  47. Stone D. E., Craig E. A. Self-regulation of 70-kilodalton heat shock proteins in Saccharomyces cerevisiae. Mol Cell Biol. 1990 Apr;10(4):1622–1632. doi: 10.1128/mcb.10.4.1622. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. Straus D. B., Walter W. A., Gross C. A. The activity of sigma 32 is reduced under conditions of excess heat shock protein production in Escherichia coli. Genes Dev. 1989 Dec;3(12A):2003–2010. doi: 10.1101/gad.3.12a.2003. [DOI] [PubMed] [Google Scholar]
  49. Straus D. B., Walter W. A., Gross C. A. The heat shock response of E. coli is regulated by changes in the concentration of sigma 32. Nature. 1987 Sep 24;329(6137):348–351. doi: 10.1038/329348a0. [DOI] [PubMed] [Google Scholar]
  50. Straus D., Walter W., Gross C. A. DnaK, DnaJ, and GrpE heat shock proteins negatively regulate heat shock gene expression by controlling the synthesis and stability of sigma 32. Genes Dev. 1990 Dec;4(12A):2202–2209. doi: 10.1101/gad.4.12a.2202. [DOI] [PubMed] [Google Scholar]
  51. Tilly K., McKittrick N., Zylicz M., Georgopoulos C. The dnaK protein modulates the heat-shock response of Escherichia coli. Cell. 1983 Sep;34(2):641–646. doi: 10.1016/0092-8674(83)90396-3. [DOI] [PubMed] [Google Scholar]
  52. Tilly K., Spence J., Georgopoulos C. Modulation of stability of the Escherichia coli heat shock regulatory factor sigma. J Bacteriol. 1989 Mar;171(3):1585–1589. doi: 10.1128/jb.171.3.1585-1589.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  53. Westwood J. T., Clos J., Wu C. Stress-induced oligomerization and chromosomal relocalization of heat-shock factor. Nature. 1991 Oct 31;353(6347):822–827. doi: 10.1038/353822a0. [DOI] [PubMed] [Google Scholar]
  54. Zimarino V., Wilson S., Wu C. Antibody-mediated activation of Drosophila heat shock factor in vitro. Science. 1990 Aug 3;249(4968):546–549. doi: 10.1126/science.2200124. [DOI] [PubMed] [Google Scholar]
  55. Zimarino V., Wu C. Induction of sequence-specific binding of Drosophila heat shock activator protein without protein synthesis. 1987 Jun 25-Jul 1Nature. 327(6124):727–730. doi: 10.1038/327727a0. [DOI] [PubMed] [Google Scholar]

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

RESOURCES