The wing in yeast heat shock transcription factor (HSF) DNA-binding domain is required for full activity

In vitro DNA-binding assays demonstrate that the heat shock transcription factor (HSF) from the yeast Saccharomyces cerevisiae can adopt an altered conformation when stressed. This conformation, reflected in a change in electrophoretic mobility, requires that two HSF trimers be bound to DNA. Single trimers do not show this change, which appears to represent an alteration in the cooperative interactions between trimers. HSF isolated from stressed cells displays a higher propensity to adopt this altered conformation. Purified HSF can be stimulated in vitro to undergo the conformational change by elevating the temperature or by exposing HSF to superoxide anion. Mutational analysis maps a region critical for this conformational change to the flexible loop between the minimal DNA-binding domain and the flexible linker that joins the DNA-binding domain to the trimerization domain. The significance of these findings is discussed in the context of the induction of the heat shock response by ischemic stroke, hypoxia, and recovery from anoxia, all known to stimulate the production of superoxide.

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Mutational analysis of human heat-shock transcription factor 1 reveals a regulatory role for oligomerization in DNA-binding specificity

Biochemical Journal ◽

10.1042/bj20090922 ◽

2009 ◽

Vol 424 (2) ◽

pp. 253-261 ◽

Cited By ~ 16

Author(s):

Yukiko Takemori ◽

Yasuaki Enoki ◽

Noritaka Yamamoto ◽

Yo Fukai ◽

Kaori Adachi ◽

...

Keyword(s):

Gene Expression ◽

Transcription Factor ◽

Heat Shock ◽

Dna Binding ◽

Binding Domain ◽

Heat Shock Transcription Factor ◽

Yeast Cells ◽

Dna Binding Domain ◽

Regulate Gene Expression ◽

Regulate Gene

HSF (heat-shock transcription factor) trimers bind to the HSE (heat-shock element) regulatory sequence of target genes and regulate gene expression. A typical HSE consists of at least three contiguous inverted repeats of the 5-bp sequence nGAAn. Yeast HSF is able to recognize discontinuous HSEs that contain gaps in the array of the nGAAn sequence; however, hHSF1 (human HSF1) fails to recognize such sites in vitro, in yeast and in HeLa cells. In the present study, we isolated suppressors of the temperature-sensitive growth defect of hHSF1-expressing yeast cells. Intragenic suppressors contained amino acid substitutions in the DNA-binding domain of hHSF1 that enabled hHSF1 to regulate the transcription of genes containing discontinuous HSEs. The substitutions facilitated hHSF1 oligomerization, suggesting that the DNA-binding domain is important for this conformational change. Furthermore, other oligomerization-prone derivatives of hHSF1 were capable of recognizing discontinuous HSEs. These results suggest that modulation of oligomerization is important for the HSE specificity of hHSF1 and imply that hHSF1 possesses the ability to bind to and regulate gene expression via various types of HSEs in diverse cellular processes.

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Crystal structure of the DNA binding domain of the heat shock transcription factor

Science ◽

10.1126/science.8284672 ◽

1994 ◽

Vol 263 (5144) ◽

pp. 224-227 ◽

Cited By ~ 170

Author(s):

C. Harrison ◽

A. Bohm ◽

H. Nelson

Keyword(s):

Crystal Structure ◽

Transcription Factor ◽

Heat Shock ◽

Dna Binding ◽

Binding Domain ◽

Heat Shock Transcription Factor ◽

Dna Binding Domain

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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.

Molecular and Cellular Biology ◽

10.1128/mcb.14.11.7557 ◽

1994 ◽

Vol 14 (11) ◽

pp. 7557-7568 ◽

Cited By ~ 126

Author(s):

J Zuo ◽

R Baler ◽

G Dahl ◽

R Voellmy

Keyword(s):

Transcription Factor ◽

Heat Stress ◽

Heat Shock ◽

Dna Binding ◽

Hydrophobic Interactions ◽

Coiled Coil ◽

Heat Shock Transcription Factor ◽

Single Amino Acid ◽

Binding Ability ◽

Heat Shock Element

Heat stress regulation of human heat shock genes is mediated by human heat shock transcription factor hHSF1, which contains three 4-3 hydrophobic repeats (LZ1 to LZ3). In unstressed human cells (37 degrees C), hHSF1 appears to be in an inactive, monomeric state that may be maintained through intramolecular interactions stabilized by transient interaction with hsp70. Heat stress (39 to 42 degrees C) disrupts these interactions, and hHSF1 homotrimerizes and acquires heat shock element DNA-binding ability. hHSF1 expressed in Xenopus oocytes also assumes a monomeric, non-DNA-binding state and is converted to a trimeric, DNA-binding form upon exposure of the oocytes to heat shock (35 to 37 degrees C in this organism). Because endogenous HSF DNA-binding activity is low and anti-hHSF1 antibody does not recognize Xenopus HSF, we employed this system for mapping regions in hHSF1 that are required for the maintenance of the monomeric state. The results of mutagenesis analyses strongly suggest that the inactive hHSF1 monomer is stabilized by hydrophobic interactions involving all three leucine zippers which may form a triple-stranded coiled coil. Trimerization may enable the DNA-binding function of hHSF1 by facilitating cooperative binding of monomeric DNA-binding domains to the heat shock element motif. This view is supported by observations that several different LexA DNA-binding domain-hHSF1 chimeras bind to a LexA-binding site in a heat-regulated fashion, that single amino acid replacements disrupting the integrity of hydrophobic repeats render these chimeras constitutively trimeric and DNA binding, and that LexA itself binds stably to DNA only as a dimer but not as a monomer in our assays.

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