Skip to main content
PMC home page
Molecular and Cellular Biology logoLink to Molecular and Cellular Biology
. 1995 Aug;15(8):4309–4318. doi: 10.1128/mcb.15.8.4309

The carboxyl-terminal transactivation domain of heat shock factor 1 is negatively regulated and stress responsive.

Y Shi 1, P E Kroeger 1, R I Morimoto 1
PMCID: PMC230670  PMID: 7623825

Abstract

We have characterized a stress-responsive transcriptional activation domain of mouse heat shock factor 1 (HSF1) by using chimeric GAL4-HSF1 fusion proteins. Fusion of the GAL4 DNA-binding domain to residues 124 to 503 of HSF1 results in a chimeric factor that binds DNA yet lacks any transcriptional activity. Transactivation is acquired upon exposure to heat shock or by deletion of a negative regulatory domain including part of the DNA-binding-domain-proximal leucine zippers. Analysis of a collection of GAL4-HSF1 deletion mutants revealed the minimal region for the constitutive transcriptional activator to map within the extreme carboxyl-terminal 108 amino acids, corresponding to a region rich in acidic and hydrophobic residues. Loss of residues 395 to 425 or 451 to 503, which are located at either end of this activation domain, severely diminished activity, indicating that the entire domain is required for transactivation. The minimal activation domain of HSF1 also confers enhanced transcriptional response to heat shock or cadmium treatment. These results demonstrate that the transcriptional activation domain of HSF1 is negatively regulated and that the signal for stress induction is mediated by interactions between the amino-terminal negative regulator and the carboxyl-terminal transcriptional activation domain.

Full Text

The Full Text of this article is available as a PDF (626.0 KB).

Selected References

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

  1. Abate C., Luk D., Curran T. Transcriptional regulation by Fos and Jun in vitro: interaction among multiple activator and regulatory domains. Mol Cell Biol. 1991 Jul;11(7):3624–3632. doi: 10.1128/mcb.11.7.3624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. 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]
  3. 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]
  4. Abravaya K., Phillips B., Morimoto R. I. Heat shock-induced interactions of heat shock transcription factor and the human hsp70 promoter examined by in vivo footprinting. Mol Cell Biol. 1991 Jan;11(1):586–592. doi: 10.1128/mcb.11.1.586. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. 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]
  6. Baler R., Dahl G., Voellmy R. Activation of human heat shock genes is accompanied by oligomerization, modification, and rapid translocation of heat shock transcription factor HSF1. Mol Cell Biol. 1993 Apr;13(4):2486–2496. doi: 10.1128/mcb.13.4.2486. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. 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]
  8. Becker P. B., Wu C. Cell-free system for assembly of transcriptionally repressed chromatin from Drosophila embryos. Mol Cell Biol. 1992 May;12(5):2241–2249. doi: 10.1128/mcb.12.5.2241. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Benjamin I. J., Kröger B., Williams R. S. Activation of the heat shock transcription factor by hypoxia in mammalian cells. Proc Natl Acad Sci U S A. 1990 Aug;87(16):6263–6267. doi: 10.1073/pnas.87.16.6263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Blake M. J., Udelsman R., Feulner G. J., Norton D. D., Holbrook N. J. Stress-induced heat shock protein 70 expression in adrenal cortex: an adrenocorticotropic hormone-sensitive, age-dependent response. Proc Natl Acad Sci U S A. 1991 Nov 1;88(21):9873–9877. doi: 10.1073/pnas.88.21.9873. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Bonner J. J., Heyward S., Fackenthal D. L. Temperature-dependent regulation of a heterologous transcriptional activation domain fused to yeast heat shock transcription factor. Mol Cell Biol. 1992 Mar;12(3):1021–1030. doi: 10.1128/mcb.12.3.1021. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Boorstein W. R., Craig E. A. Structure and regulation of the SSA4 HSP70 gene of Saccharomyces cerevisiae. J Biol Chem. 1990 Nov 5;265(31):18912–18921. [PubMed] [Google Scholar]
  13. 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]
  14. Chen Y., Barlev N. A., Westergaard O., Jakobsen B. K. Identification of the C-terminal activator domain in yeast heat shock factor: independent control of transient and sustained transcriptional activity. EMBO J. 1993 Dec 15;12(13):5007–5018. doi: 10.1002/j.1460-2075.1993.tb06194.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Chomczynski P., Sacchi N. Single-step method of RNA isolation by acid guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem. 1987 Apr;162(1):156–159. doi: 10.1006/abio.1987.9999. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Ferris D. K., Harel-Bellan A., Morimoto R. I., Welch W. J., Farrar W. L. Mitogen and lymphokine stimulation of heat shock proteins in T lymphocytes. Proc Natl Acad Sci U S A. 1988 Jun;85(11):3850–3854. doi: 10.1073/pnas.85.11.3850. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Gallo G. J., Schuetz T. J., Kingston R. E. Regulation of heat shock factor in Schizosaccharomyces pombe more closely resembles regulation in mammals than in Saccharomyces cerevisiae. Mol Cell Biol. 1991 Jan;11(1):281–288. doi: 10.1128/mcb.11.1.281. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. 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]
  20. Greene J. M., Kingston R. E. TATA-dependent and TATA-independent function of the basal and heat shock elements of a human hsp70 promoter. Mol Cell Biol. 1990 Apr;10(4):1319–1328. doi: 10.1128/mcb.10.4.1319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Greene J. M., Larin Z., Taylor I. C., Prentice H., Gwinn K. A., Kingston R. E. Multiple basal elements of a human hsp70 promoter function differently in human and rodent cell lines. Mol Cell Biol. 1987 Oct;7(10):3646–3655. doi: 10.1128/mcb.7.10.3646. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hunt C., Calderwood S. Characterization and sequence of a mouse hsp70 gene and its expression in mouse cell lines. Gene. 1990 Mar 15;87(2):199–204. doi: 10.1016/0378-1119(90)90302-8. [DOI] [PubMed] [Google Scholar]
  23. Jurivich D. A., Sistonen L., Kroes R. A., Morimoto R. I. Effect of sodium salicylate on the human heat shock response. Science. 1992 Mar 6;255(5049):1243–1245. doi: 10.1126/science.1546322. [DOI] [PubMed] [Google Scholar]
  24. Jurivich D. A., Sistonen L., Sarge K. D., Morimoto R. I. Arachidonate is a potent modulator of human heat shock gene transcription. Proc Natl Acad Sci U S A. 1994 Mar 15;91(6):2280–2284. doi: 10.1073/pnas.91.6.2280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. 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]
  26. 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]
  27. Lis J., Wu C. Protein traffic on the heat shock promoter: parking, stalling, and trucking along. Cell. 1993 Jul 16;74(1):1–4. doi: 10.1016/0092-8674(93)90286-y. [DOI] [PubMed] [Google Scholar]
  28. Lum L. S., Sultzman L. A., Kaufman R. J., Linzer D. I., Wu B. J. A cloned human CCAAT-box-binding factor stimulates transcription from the human hsp70 promoter. Mol Cell Biol. 1990 Dec;10(12):6709–6717. doi: 10.1128/mcb.10.12.6709. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Ma J., Ptashne M. Deletion analysis of GAL4 defines two transcriptional activating segments. Cell. 1987 Mar 13;48(5):847–853. doi: 10.1016/0092-8674(87)90081-x. [DOI] [PubMed] [Google Scholar]
  30. Mathur S. K., Sistonen L., Brown I. R., Murphy S. P., Sarge K. D., Morimoto R. I. Deficient induction of human hsp70 heat shock gene transcription in Y79 retinoblastoma cells despite activation of heat shock factor 1. Proc Natl Acad Sci U S A. 1994 Aug 30;91(18):8695–8699. doi: 10.1073/pnas.91.18.8695. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. Morgan W. D. Transcription factor Sp1 binds to and activates a human hsp70 gene promoter. Mol Cell Biol. 1989 Sep;9(9):4099–4104. doi: 10.1128/mcb.9.9.4099. [DOI] [PMC free article] [PubMed] [Google Scholar]
  32. Morgan W. D., Williams G. T., Morimoto R. I., Greene J., Kingston R. E., Tjian R. Two transcriptional activators, CCAAT-box-binding transcription factor and heat shock transcription factor, interact with a human hsp70 gene promoter. Mol Cell Biol. 1987 Mar;7(3):1129–1138. doi: 10.1128/mcb.7.3.1129. [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Morimoto R. I. Cells in stress: transcriptional activation of heat shock genes. Science. 1993 Mar 5;259(5100):1409–1410. doi: 10.1126/science.8451637. [DOI] [PubMed] [Google Scholar]
  34. 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]
  35. 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]
  36. Nakai A., Morimoto R. I. Characterization of a novel chicken heat shock transcription factor, heat shock factor 3, suggests a new regulatory pathway. Mol Cell Biol. 1993 Apr;13(4):1983–1997. doi: 10.1128/mcb.13.4.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. 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]
  38. Peteranderl R., Nelson H. C. Trimerization of the heat shock transcription factor by a triple-stranded alpha-helical coiled-coil. Biochemistry. 1992 Dec 8;31(48):12272–12276. doi: 10.1021/bi00163a042. [DOI] [PubMed] [Google Scholar]
  39. Phillips B., Morimoto R. I. Transcriptional regulation of human hsp70 genes: relationship between cell growth, differentiation, virus infection, and the stress response. Results Probl Cell Differ. 1991;17:167–187. doi: 10.1007/978-3-540-46712-0_12. [DOI] [PubMed] [Google Scholar]
  40. 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]
  41. 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]
  42. 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]
  43. 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]
  44. Scharf K. D., Rose S., Zott W., Schöffl F., Nover L., Schöff F. Three tomato genes code for heat stress transcription factors with a region of remarkable homology to the DNA-binding domain of the yeast HSF. EMBO J. 1990 Dec;9(13):4495–4501. doi: 10.1002/j.1460-2075.1990.tb07900.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. 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]
  46. Sistonen L., Sarge K. D., Morimoto R. I. Human heat shock factors 1 and 2 are differentially activated and can synergistically induce hsp70 gene transcription. Mol Cell Biol. 1994 Mar;14(3):2087–2099. doi: 10.1128/mcb.14.3.2087. [DOI] [PMC free article] [PubMed] [Google Scholar]
  47. Sistonen L., Sarge K. D., Phillips B., Abravaya K., Morimoto R. I. Activation of heat shock factor 2 during hemin-induced differentiation of human erythroleukemia cells. Mol Cell Biol. 1992 Sep;12(9):4104–4111. doi: 10.1128/mcb.12.9.4104. [DOI] [PMC free article] [PubMed] [Google Scholar]
  48. 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]
  49. 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]
  50. 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]
  51. Sorger P. K. Yeast heat shock factor contains separable transient and sustained response transcriptional activators. Cell. 1990 Aug 24;62(4):793–805. doi: 10.1016/0092-8674(90)90123-v. [DOI] [PubMed] [Google Scholar]
  52. 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]
  53. Taylor I. C., Workman J. L., Schuetz T. J., Kingston R. E. Facilitated binding of GAL4 and heat shock factor to nucleosomal templates: differential function of DNA-binding domains. Genes Dev. 1991 Jul;5(7):1285–1298. doi: 10.1101/gad.5.7.1285. [DOI] [PubMed] [Google Scholar]
  54. Theodorakis N. G., Zand D. J., Kotzbauer P. T., Williams G. T., Morimoto R. I. Hemin-induced transcriptional activation of the HSP70 gene during erythroid maturation in K562 cells is due to a heat shock factor-mediated stress response. Mol Cell Biol. 1989 Aug;9(8):3166–3173. doi: 10.1128/mcb.9.8.3166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  55. Treuter E., Nover L., Ohme K., Scharf K. D. Promoter specificity and deletion analysis of three heat stress transcription factors of tomato. Mol Gen Genet. 1993 Jul;240(1):113–125. doi: 10.1007/BF00276890. [DOI] [PubMed] [Google Scholar]
  56. Westwood J. T., Wu C. Activation of Drosophila heat shock factor: conformational change associated with a monomer-to-trimer transition. Mol Cell Biol. 1993 Jun;13(6):3481–3486. doi: 10.1128/mcb.13.6.3481. [DOI] [PMC free article] [PubMed] [Google Scholar]
  57. Wiederrecht G., Seto D., Parker C. S. Isolation of the gene encoding the S. cerevisiae heat shock transcription factor. Cell. 1988 Sep 9;54(6):841–853. doi: 10.1016/s0092-8674(88)91197-x. [DOI] [PubMed] [Google Scholar]
  58. Williams G. T., McClanahan T. K., Morimoto R. I. E1a transactivation of the human HSP70 promoter is mediated through the basal transcriptional complex. Mol Cell Biol. 1989 Jun;9(6):2574–2587. doi: 10.1128/mcb.9.6.2574. [DOI] [PMC free article] [PubMed] [Google Scholar]
  59. Williams G. T., Morimoto R. I. Maximal stress-induced transcription from the human HSP70 promoter requires interactions with the basal promoter elements independent of rotational alignment. Mol Cell Biol. 1990 Jun;10(6):3125–3136. doi: 10.1128/mcb.10.6.3125. [DOI] [PMC free article] [PubMed] [Google Scholar]
  60. 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]
  61. 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]
  62. Xiao H., Lis J. T. Heat shock and developmental regulation of the Drosophila melanogaster hsp83 gene. Mol Cell Biol. 1989 Apr;9(4):1746–1753. doi: 10.1128/mcb.9.4.1746. [DOI] [PMC free article] [PubMed] [Google Scholar]

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

RESOURCES