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Structure of human heat-shock transcription factor 1 in complex with DNA

Nature Structural & Molecular Biology volume 23, pages 140–146 (2016)Cite this article

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Abstract

Heat-shock transcription factor 1 (HSF1) has a central role in mediating the protective response to protein conformational stresses in eukaryotes. HSF1 consists of an N-terminal DNA-binding domain (DBD), a coiled-coil oligomerization domain, a regulatory domain and a transactivation domain. Upon stress, HSF1 trimerizes via its coiled-coil domain and binds to the promoters of heat shock protein–encoding genes. Here, we present cocrystal structures of the human HSF1 DBD in complex with cognate DNA. A comparative analysis of the HSF1 paralog Skn7 from Chaetomium thermophilum showed that single amino acid changes in the DBD can switch DNA binding specificity, thus revealing the structural basis for the interaction of HSF1 with cognate DNA. We used a crystal structure of the coiled-coil domain of C. thermophilum Skn7 to develop a model of the active human HSF1 trimer in which HSF1 embraces the heat-shock-element DNA.

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Figure 1: Interaction of HsHSF1 with cognate DNA.
Figure 2: Symmetrical interactions between the HsDBDs in the complex with the tail-to-tail HSE.
Figure 3: Characterization of the HSF homolog Skn7 of C. thermophilum.
Figure 4: Interaction of CtSkn7 with DNA.
Figure 5: Oligomerization of CtSkn7 via the coiled-coil domain.
Figure 6: Structural model for HsHSF1(13–182) bound to an HSE with three inverted GAA repeats.

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References

  1. Balch, W.E., Morimoto, R.I., Dillin, A. & Kelly, J.W. Adapting proteostasis for disease intervention. Science 319, 916–919 (2008).

    Article  CAS  Google Scholar 

  2. Kim, Y.E., Hipp, M.S., Bracher, A., Hayer-Hartl, M. & Hartl, F.U. Molecular chaperone functions in protein folding and proteostasis. Annu. Rev. Biochem. 82, 323–355 (2013).

    Article  CAS  Google Scholar 

  3. Åkerfelt, M., Morimoto, R.I. & Sistonen, L. Heat shock factors: integrators of cell stress, development and lifespan. Nat. Rev. Mol. Cell Biol. 11, 545–555 (2010).

    Article  Google Scholar 

  4. Vabulas, R.M., Raychaudhuri, S., Hayer-Hartl, M. & Hartl, F.U. Protein folding in the cytoplasm and the heat shock response. Cold Spring Harb. Perspect. Biol. 2, a004390 (2010).

    Article  CAS  Google Scholar 

  5. Anckar, J. & Sistonen, L. Regulation of HSF1 function in the heat stress response: implications in aging and disease. Annu. Rev. Biochem. 80, 1089–1115 (2011).

    Article  CAS  Google Scholar 

  6. Morimoto, R.I. The heat shock response: systems biology of proteotoxic stress in aging and disease. Cold Spring Harb. Symp. Quant. Biol. 76, 91–99 (2011).

    Article  CAS  Google Scholar 

  7. Pelham, H.R. A regulatory upstream promoter element in the Drosophila hsp 70 heat-shock gene. Cell 30, 517–528 (1982).

    Article  CAS  Google Scholar 

  8. Amin, J., Ananthan, J. & Voellmy, R. Key features of heat shock regulatory elements. Mol. Cell. Biol. 8, 3761–3769 (1988).

    Article  CAS  Google Scholar 

  9. Xiao, H. & Lis, J.T. Germline transformation used to define key features of heat-shock response elements. Science 239, 1139–1142 (1988).

    Article  CAS  Google Scholar 

  10. Wu, C. Heat shock transcription factors: structure and regulation. Annu. Rev. Cell Dev. Biol. 11, 441–469 (1995).

    Article  CAS  Google Scholar 

  11. Pattaramanon, N., Sangha, N. & Gafni, A. The carboxy-terminal domain of heat-shock factor 1 is largely unfolded but can be induced to collapse into a compact, partially structured state. Biochemistry 46, 3405–3415 (2007).

    Article  CAS  Google Scholar 

  12. Harrison, C.J., Bohm, A.A. & Nelson, H.C. Crystal structure of the DNA binding domain of the heat shock transcription factor. Science 263, 224–227 (1994).

    Article  CAS  Google Scholar 

  13. Vuister, G.W. et al. Solution structure of the DNA-binding domain of Drosophila heat shock transcription factor. Nat. Struct. Biol. 1, 605–614 (1994).

    Article  CAS  Google Scholar 

  14. Liu, G. et al. Solution NMR structure of heat shock factor protein 1 DNA binding domain from Homo sapiens, Northeast Structural Genomics Consortium Target HR3023C (2011).

  15. Littlefield, O. & Nelson, H.C. A new use for the 'wing' of the 'winged' helix-turn-helix motif in the HSF–DNA cocrystal. Nat. Struct. Biol. 6, 464–470 (1999).

    Article  CAS  Google Scholar 

  16. 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. 13, 2486–2496 (1993).

    Article  CAS  Google Scholar 

  17. 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. 13, 1392–1407 (1993).

    Article  CAS  Google Scholar 

  18. Xiao, X. et al. HSF1 is required for extra-embryonic development, postnatal growth and protection during inflammatory responses in mice. EMBO J. 18, 5943–5952 (1999).

    Article  CAS  Google Scholar 

  19. Mendillo, M.L. et al. HSF1 drives a transcriptional program distinct from heat shock to support highly malignant human cancers. Cell 150, 549–562 (2012).

    Article  CAS  Google Scholar 

  20. Santagata, S. et al. Tight coordination of protein translation and HSF1 activation supports the anabolic malignant state. Science 341, 1238303 (2013).

    Article  Google Scholar 

  21. Dai, C., Whitesell, L., Rogers, A.B. & Lindquist, S. Heat shock factor 1 is a powerful multifaceted modifier of carcinogenesis. Cell 130, 1005–1018 (2007).

    Article  CAS  Google Scholar 

  22. Scherz-Shouval, R. et al. The reprogramming of tumor stroma by HSF1 is a potent enabler of malignancy. Cell 158, 564–578 (2014).

    Article  CAS  Google Scholar 

  23. Mercier, P.A., Winegarden, N.A. & Westwood, J.T. Human heat shock factor 1 is predominantly a nuclear protein before and after heat stress. J. Cell Sci. 112, 2765–2774 (1999).

    CAS  PubMed  Google Scholar 

  24. 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 259, 230–234 (1993).

    Article  CAS  Google Scholar 

  25. Zuo, J., Baler, R., Dahl, G. & Voellmy, R. Activation of the DNA-binding ability of human heat shock transcription factor 1 may involve the transition from an intramolecular to an intermolecular triple-stranded coiled-coil structure. Mol. Cell. Biol. 14, 7557–7568 (1994).

    Article  CAS  Google Scholar 

  26. Neef, D.W., Jaeger, A.M. & Thiele, D.J. Genetic selection for constitutively trimerized human HSF1 mutants identifies a role for coiled-coil motifs in DNA binding. G3 (Bethesda) 3, 1315–1324 (2013).

    Article  Google Scholar 

  27. 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. 6, 1153–1164 (1992).

    Article  CAS  Google Scholar 

  28. 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. 268, 1479–1487 (1993).

    CAS  PubMed  Google Scholar 

  29. Zou, J., Guo, Y., Guettouche, T., Smith, D.F. & Voellmy, R. Repression of heat shock transcription factor HSF1 activation by HSP90 (HSP90 complex) that forms a stress-sensitive complex with HSF1. Cell 94, 471–480 (1998).

    Article  CAS  Google Scholar 

  30. Neef, D.W. et al. A direct regulatory interaction between chaperonin TRiC and stress-responsive transcription factor HSF1. Cell Rep. 9, 955–966 (2014).

    Article  CAS  Google Scholar 

  31. Shi, Y., Mosser, D.D. & Morimoto, R.I. Molecular chaperones as HSF1-specific transcriptional repressors. Genes Dev. 12, 654–666 (1998).

    Article  CAS  Google Scholar 

  32. Morimoto, R.I. Dynamic remodeling of transcription complexes by molecular chaperones. Cell 110, 281–284 (2002).

    Article  CAS  Google Scholar 

  33. Larson, J.S., Schuetz, T.J. & Kingston, R.E. In vitro activation of purified human heat shock factor by heat. Biochemistry 34, 1902–1911 (1995).

    Article  CAS  Google Scholar 

  34. Kline, M.P. & Morimoto, R.I. Repression of the heat shock factor 1 transcriptional activation domain is modulated by constitutive phosphorylation. Mol. Cell. Biol. 17, 2107–2115 (1997).

    Article  CAS  Google Scholar 

  35. Hietakangas, V. et al. Phosphorylation of serine 303 is a prerequisite for the stress-inducible SUMO modification of heat shock factor 1. Mol. Cell. Biol. 23, 2953–2968 (2003).

    Article  CAS  Google Scholar 

  36. Westerheide, S.D., Anckar, J., Stevens, S.M. Jr., Sistonen, L. & Morimoto, R.I. Stress-inducible regulation of heat shock factor 1 by the deacetylase SIRT1. Science 323, 1063–1066 (2009).

    Article  CAS  Google Scholar 

  37. Raychaudhuri, S. et al. Interplay of acetyltransferase EP300 and the proteasome system in regulating heat shock transcription factor 1. Cell 156, 975–985 (2014).

    Article  CAS  Google Scholar 

  38. Grady, D.L. et al. Highly conserved repetitive DNA sequences are present at human centromeres. Proc. Natl. Acad. Sci. USA 89, 1695–1699 (1992).

    Article  CAS  Google Scholar 

  39. Denegri, M. et al. Human chromosomes 9, 12, and 15 contain the nucleation sites of stress-induced nuclear bodies. Mol. Biol. Cell 13, 2069–2079 (2002).

    Article  CAS  Google Scholar 

  40. Jolly, C. et al. In vivo binding of active heat shock transcription factor 1 to human chromosome 9 heterochromatin during stress. J. Cell Biol. 156, 775–781 (2002).

    Article  CAS  Google Scholar 

  41. Jolly, C. et al. Stress-induced transcription of satellite III repeats. J. Cell Biol. 164, 25–33 (2004).

    Article  CAS  Google Scholar 

  42. Metz, A., Soret, J., Vourc'h, C., Tazi, J. & Jolly, C. A key role for stress-induced satellite III transcripts in the relocalization of splicing factors into nuclear stress granules. J. Cell Sci. 117, 4551–4558 (2004).

    Article  CAS  Google Scholar 

  43. Flick, K.E., Gonzalez, L. Jr., Harrison, C.J. & Nelson, H.C. Yeast heat shock transcription factor contains a flexible linker between the DNA-binding and trimerization domains: implications for DNA binding by trimeric proteins. J. Biol. Chem. 269, 12475–12481 (1994).

    CAS  PubMed  Google Scholar 

  44. Zelin, E., Zhang, Y., Toogun, O.A., Zhong, S. & Freeman, B.C. The p23 molecular chaperone and GCN5 acetylase jointly modulate protein-DNA dynamics and open chromatin status. Mol. Cell 48, 459–470 (2012).

    Article  CAS  Google Scholar 

  45. Ahn, S.G. & Thiele, D.J. Redox regulation of mammalian heat shock factor 1 is essential for Hsp gene activation and protection from stress. Genes Dev. 17, 516–528 (2003).

    Article  CAS  Google Scholar 

  46. Lu, M. et al. Two distinct disulfide bonds formed in human heat shock transcription factor 1 act in opposition to regulate its DNA binding activity. Biochemistry 47, 6007–6015 (2008).

    Article  CAS  Google Scholar 

  47. Xiao, H., Perisic, O. & Lis, J.T. Cooperative binding of Drosophila heat shock factor to arrays of a conserved 5 bp unit. Cell 64, 585–593 (1991).

    Article  CAS  Google Scholar 

  48. Sorger, P.K. & Pelham, H.R. Yeast heat shock factor is an essential DNA-binding protein that exhibits temperature-dependent phosphorylation. Cell 54, 855–864 (1988).

    Article  CAS  Google Scholar 

  49. Wiederrecht, G., Seto, D. & Parker, C.S. Isolation of the gene encoding the S. cerevisiae heat shock transcription factor. Cell 54, 841–853 (1988).

    Article  CAS  Google Scholar 

  50. Fassler, J.S. & West, A.H. Fungal Skn7 stress responses and their relationship to virulence. Eukaryot. Cell 10, 156–167 (2011).

    Article  CAS  Google Scholar 

  51. Li, S. et al. The yeast histidine protein kinase, Sln1p, mediates phosphotransfer to two response regulators, Ssk1p and Skn7p. EMBO J. 17, 6952–6962 (1998).

    Article  CAS  Google Scholar 

  52. Morgan, B.A. et al. The Skn7 response regulator controls gene expression in the oxidative stress response of the budding yeast Saccharomyces cerevisiae. EMBO J. 16, 1035–1044 (1997).

    Article  CAS  Google Scholar 

  53. Li, S. et al. The eukaryotic two-component histidine kinase Sln1p regulates OCH1 via the transcription factor, Skn7p. Mol. Biol. Cell 13, 412–424 (2002).

    Article  CAS  Google Scholar 

  54. Peteranderl, R. & Nelson, H.C. Trimerization of the heat shock transcription factor by a triple-stranded alpha-helical coiled-coil. Biochemistry 31, 12272–12276 (1992).

    Article  CAS  Google Scholar 

  55. Sandqvist, A. et al. Heterotrimerization of heat-shock factors 1 and 2 provides a transcriptional switch in response to distinct stimuli. Mol. Biol. Cell 20, 1340–1347 (2009).

    Article  CAS  Google Scholar 

  56. Neef, D.W., Jaeger, A.M. & Thiele, D.J. Heat shock transcription factor 1 as a therapeutic target in neurodegenerative diseases. Nat. Rev. Drug Discov. 10, 930–944 (2011).

    Article  CAS  Google Scholar 

  57. Alarcon, S.V. et al. Tumor-intrinsic and tumor-extrinsic factors impacting hsp90- targeted therapy. Curr. Mol. Med. 12, 1125–1141 (2012).

    Article  CAS  Google Scholar 

  58. Calamini, B. & Morimoto, R.I. Protein homeostasis as a therapeutic target for diseases of protein conformation. Curr. Top. Med. Chem. 12, 2623–2640 (2012).

    Article  CAS  Google Scholar 

  59. Treisman, J., Gönczy, P., Vashishtha, M., Harris, E. & Desplan, C. A single amino acid can determine the DNA binding specificity of homeodomain proteins. Cell 59, 553–562 (1989).

    Article  CAS  Google Scholar 

  60. Tucker-Kellogg, L. et al. Engrailed (Gln50Lys) homeodomain-DNA complex at 1.9 Å resolution: structural basis for enhanced affinity and altered specificity. Structure 5, 1047–1054 (1997).

    Article  CAS  Google Scholar 

  61. Catanzariti, A.M., Soboleva, T.A., Jans, D.A., Board, P.G. & Baker, R.T. An efficient system for high-level expression and easy purification of authentic recombinant proteins. Protein Sci. 13, 1331–1339 (2004).

    Article  CAS  Google Scholar 

  62. Jaeger, A.M., Makley, L.N., Gestwicki, J.E. & Thiele, D.J. Genomic heat shock element sequences drive cooperative human heat shock factor 1 DNA binding and selectivity. J. Biol. Chem. 289, 30459–30469 (2014).

    Article  CAS  Google Scholar 

  63. Wyatt, P.J. Light scattering and the absolute characterization of macromolecules. Anal. Chim. Acta 272, 1–40 (1993).

    Article  CAS  Google Scholar 

  64. Kabsch, W. XDS. Acta Crystallogr. D Biol. Crystallogr. 66, 125–132 (2010).

    Article  CAS  Google Scholar 

  65. Evans, P. Scaling and assessment of data quality. Acta Crystallogr. D Biol. Crystallogr. 62, 72–82 (2006).

    Article  Google Scholar 

  66. Evans, P.R. Scala. Joint CCP4 ESF-EACBM Newsl. 33, 22–24 (1997).

    Google Scholar 

  67. Evans, P.R. & Murshudov, G.N. How good are my data and what is the resolution? Acta Crystallogr. D Biol. Crystallogr. 69, 1204–1214 (2013).

    Article  CAS  Google Scholar 

  68. French, G. & Wilson, K. On the treatment of negative intensity observations. Acta Crystallogr. A 34, 517–525 (1978).

    Article  Google Scholar 

  69. Collaborative Computational Project, Number 4. The CCP4 suite: programs for protein crystallography. Acta Crystallogr. D Biol. Crystallogr. 50, 760–763 (1994).

  70. Pape, T. & Schneider, T.R. HKL2MAP: a graphical user interface for phasing with SHELX programs. J. Appl. Crystallogr. 37, 843–844 (2004).

    Article  CAS  Google Scholar 

  71. Sheldrick, G.M. Experimental phasing with SHELXC/D/E: combining chain tracing with density modification. Acta Crystallogr. D Biol. Crystallogr. 66, 479–485 (2010).

    Article  CAS  Google Scholar 

  72. Langer, G., Cohen, S.X., Lamzin, V.S. & Perrakis, A. Automated macromolecular model building for X-ray crystallography using ARP/wARP version 7. Nat. Protoc. 3, 1171–1179 (2008).

    Article  CAS  Google Scholar 

  73. Vagin, A.A. & Isupov, M.N. Spherically averaged phased translation function and its application to the search for molecules and fragments in electron-density maps. Acta Crystallogr. D Biol. Crystallogr. 57, 1451–1456 (2001).

    Article  CAS  Google Scholar 

  74. Emsley, P. & Cowtan, K. Coot: model-building tools for molecular graphics. Acta Crystallogr. D Biol. Crystallogr. 60, 2126–2132 (2004).

    Article  Google Scholar 

  75. Murshudov, G.N., Vagin, A.A. & Dodson, E.J. Refinement of macromolecular structures by the maximum-likelihood method. Acta Crystallogr. D Biol. Crystallogr. 53, 240–255 (1997).

    Article  CAS  Google Scholar 

  76. Kleywegt, G.T. & Jones, T.A. A super position. Joint CCP4 ESF-EACBM Newsl. 31, 9–14 (1994).

    Google Scholar 

  77. Sali, A. & Blundell, T.L. Comparative protein modelling by satisfaction of spatial restraints. J. Mol. Biol. 234, 779–815 (1993).

    Article  CAS  Google Scholar 

  78. Fiser, A., Do, R.K. & Sali, A. Modeling of loops in protein structures. Protein Sci. 9, 1753–1773 (2000).

    Article  CAS  Google Scholar 

  79. Fiser, A. & Sali, A. ModLoop: automated modeling of loops in protein structures. Bioinformatics 19, 2500–2501 (2003).

    Article  CAS  Google Scholar 

  80. Pettersen, E.F. et al. UCSF Chimera: a visualization system for exploratory research and analysis. J. Comput. Chem. 25, 1605–1612 (2004).

    Article  CAS  Google Scholar 

  81. Gouet, P., Courcelle, E., Stuart, D.I. & Métoz, F. ESPript: analysis of multiple sequence alignments in PostScript. Bioinformatics 15, 305–308 (1999).

    Article  CAS  Google Scholar 

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Acknowledgements

We thank R. Körner for MS analysis, R. Lange and A. Jungclaus for assistance with protein purification, and the staff at the Core and Crystallization Facilities at the Max Planck Institute of Biochemistry and at the European Synchrotron Radiation Facility (ESRF) in Grenoble, France, for their excellent services. J.V. is supported by a Rudolf Haas Fellowship from the Jung Foundation for Science and Research.

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  1. Department of Cellular Biochemistry, Max Planck Institute of Biochemistry, Martinsried, Germany

    Tobias Neudegger, Jacob Verghese, Manajit Hayer-Hartl, F Ulrich Hartl & Andreas Bracher

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  1. Tobias Neudegger
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Contributions

T.N. performed the biochemical and functional analysis and obtained the crystals. A.B. and T.N. solved the crystal structures, J.V. performed the cell culture experiments, and M.H.-H. collected SEC-MALS measurements. A.B. and F.U.H. supervised the experimental design and data interpretation. All authors contributed to experimental design, data analyses and manuscript preparation.

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Correspondence to F Ulrich Hartl or Andreas Bracher.

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Integrated supplementary information

Supplementary Figure 1 The DNA-binding domain of HsHSF1 (HsDBD).

(a-c) Superposition of the HsDBD structure in the HsDBD–HSE complex with the DBDs in the HsDBD–SatIII complex (a), with the NMR structure of the free HsDBD (PDB 2LDU, Liu, G. et al., 2011) (b) and with the KlDBD–HSE complex (PDB 3HTS, Littlefield, O. & Nelson, H.C. Nat. Struct. Biol. 6, 464-470, 1999) (c). Cα traces are shown. (d) Interface between the C-terminal 20 residues and the main body of the HsDBD. The protein backbone of HsDBD from the HsDBD–SatIII complex is shown in ribbon representation; residues 13-100 are additionally represented as molecular surface. The hydrophobic interface residues in the C-terminal segment are highlighted in stick representation. The hydrogen bond contact to helix α3 is shown as a dotted line.

Supplementary Figure 2 DBD-DBD interactions in HSF1–DNA complexes.

(a, b) The KlDBD–HSE complex crystal structure (pdb 3HTS) viewed along the two-fold symmetry axis (a), with the molecular contacts in the DBD–DBD interface highlighted in (b). Protein is shown in ribbon representation; DNA is shown in surface representation. Prominent sidechains are shown in stick representation; hydrogen bonds are represented by dotted lines. (c) Interactions between two asymmetric units along the DNA stacks in the crystal lattice of the HsDBD–SatIII complex. The duplexes are rotated out of register. Furthermore, two additional nucleotides were included into the DNA, shifting the DBDs ~6.8 Å further apart. (d, e) Potential interaction site of the wing domain in head-to-tail contacts. A view along the DNA is shown. A hydrophobic groove is situated between helices 2 and 3 (indicated by an arrow). Residues lining the groove are highlighted in stick representation. Panel e shows a surface representation with positive and negative charges is blue and red, respectively. Hydrophobic sidechains are shown in yellow.

Supplementary Figure 3 Sequence alignment of the conserved segment of HsHSF1, CtHSF1 and CtSkn7.

Secondary structure elements are indicated above and below the sequences. Similar residues are shown in red and identical residues in white using bold lettering on red background. Blue frames indicate homologous regions. The downward arrowhead designates the residue critical for Skn7 DNA-binding properties. Asterisks denote acetylation sites in HsHSF1; red ovals designate putative hydrophobic layer residues in the HsHSF1 coiled-coil.

Supplementary Figure 4 Interaction of CtSkn7 with DNA.

(a) Superposition of the DBDs from HsDBD–HSE and CtDBD–HSE complexes. (b) Superposition of the DBD pairs from CtDBD–HSE and CtDBD–SSRE structures. (c) Sequence alignment of representative Skn7 and HSF1 sequences, showing the systematic sequence divergence at residue 100 (CtSkn7 numbering). (d) Close-up of the interactions of CtDBD with the GGC triplet in SSRE. Sidechains of conserved residues are shown in stick representation. Hydrogen bonds are represented by dotted lines. The C-terminal tail of the DBD was omitted for clarity. (e, f) Omit density for the base-specific interactions in the CtDBD–SSRE complex. Panels e and f show the interactions of helix 3 with the GCC and GGC motifs, respectively (same orientation as in Fig. 4c). For calculating omit density, CtDBD residues 95-101 were deleted from the model, 0.2 Å random shifts applied to the coordinates with PDBSET and B-factors set to 20.00 Å2 to reduce model bias. These modified models were re-refined with Refmac5. Weighted 2Fo-Fc density maps at 1.0 sigma are shown for the missing segments, superposed on the complete model. Water atoms and the C-terminal tail residues were deleted for clarity. (g) Close-up of the interactions of CtDBD with the GAA triplet in HSE.

Supplementary Figure 5 Biophysical characterization of CtSkn7 and the chimeric HsHSF1(CtHR-A/B) protein.

(a, b) Limited proteolysis of CtSkn7(40–220) in absence (a) and presence (b) of HSE DNA duplexes. CtSkn7(40–220) at 11 μM was treated with increasing amounts of proteinase K (PK) (0, 0.05, 0.15, 0.45, 1.35, 4.05, 12.15 and 36.45 μg ml-1) for 1 h on ice. The reaction was terminated by addition of 4 mM phenylmethylsulfonyl fluoride. The products were analyzed by SDS-PAGE and Coomassie staining. (c, d) SEC-MALS analysis of CtSkn7(160–209) (c) and HsHSF1(CtHR-A/B) (d). Data show measurements of ~100 μg protein. The calculated molar mass is indicated. The theoretical molar masses of CtSkn7(160–209) dimers and trimers are ~11.4 and ~17.1 kDa, respectively. The average observed mass of ~14.5 kDa is consistent with a ~1:1 mixture of dimer and trimer. The theoretical mass for HsHSF1(CtHR-A/B) trimer is ~171.8 kDa. (e, f) Cellular locations of HsHSF1-V (e) and HsHSF1(CtHR-A/B)-V (f) in HeLa cells in absence and presence of heat shock (HS) treatment. Nuclei are stained with DAPI.

Supplementary Figure 6 Structural model for HsHSF1(13–182) bound to SatIII repeats.

Perpendicular views are shown. The C-termini point alternatingly in opposite directions; only every second HsHSF1 chain can contribute to a trimeric complex. The three HsHSF1 chains in one complex are indicated in blue, yellow and beige. The DNA is shown in stick representation. The linkers were modelled with Modeller as implemented in the ModLoop server. The helical bundle is a homology model based on the CtSkn7(160–209) trimer structure.

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Neudegger, T., Verghese, J., Hayer-Hartl, M. et al. Structure of human heat-shock transcription factor 1 in complex with DNA. Nat Struct Mol Biol 23, 140–146 (2016). https://doi.org/10.1038/nsmb.3149

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