Azadiradione ameliorates polyglutamine expansion disease in Drosophila by potentiating DNA binding activity of heat shock factor 1

The existence of multiple heat shock factor (HSF) genes in higher eukaryotes has promoted questions regarding the functions of these HSF family members, especially with respect to the stress response. To address these questions, we have used polyclonal antisera raised against mouse HSF1 and HSF2 to examine the biochemical, physical, and functional properties of these two factors in unstressed and heat-shocked mouse and human cells. We have identified HSF1 as the mediator of stress-induced heat shock gene transcription. HSF1 displays stress-induced DNA-binding activity, oligomerization, and nuclear localization, while HSF2 does not. Also, HSF1 undergoes phosphorylation in cells exposed to heat or cadmium sulfate but not in cells treated with the amino acid analog L-azetidine-2-carboxylic acid, indicating that phosphorylation of HSF1 is not essential for its activation. Interestingly, HSF1 and HSF2 overexpressed in transfected 3T3 cells both display constitutive DNA-binding activity, oligomerization, and transcriptional activity. These results demonstrate that HSF1 can be activated in the absence of physiological stress and also provide support for a model of regulation of HSF1 and HSF2 activity by a titratable negative regulatory factor.

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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

Molecular and Cellular Biology ◽

10.1128/mcb.13.3.1392-1407.1993 ◽

1993 ◽

Vol 13 (3) ◽

pp. 1392-1407

Author(s):

K D Sarge ◽

S P Murphy ◽

R I Morimoto

Keyword(s):

Heat Shock ◽

Dna Binding ◽

Gene Transcription ◽

Nuclear Localization ◽

Heat Shock Factor ◽

Binding Activity ◽

Heat Shock Gene ◽

Dna Binding Activity ◽

Shock Gene ◽

In Cells

The existence of multiple heat shock factor (HSF) genes in higher eukaryotes has promoted questions regarding the functions of these HSF family members, especially with respect to the stress response. To address these questions, we have used polyclonal antisera raised against mouse HSF1 and HSF2 to examine the biochemical, physical, and functional properties of these two factors in unstressed and heat-shocked mouse and human cells. We have identified HSF1 as the mediator of stress-induced heat shock gene transcription. HSF1 displays stress-induced DNA-binding activity, oligomerization, and nuclear localization, while HSF2 does not. Also, HSF1 undergoes phosphorylation in cells exposed to heat or cadmium sulfate but not in cells treated with the amino acid analog L-azetidine-2-carboxylic acid, indicating that phosphorylation of HSF1 is not essential for its activation. Interestingly, HSF1 and HSF2 overexpressed in transfected 3T3 cells both display constitutive DNA-binding activity, oligomerization, and transcriptional activity. These results demonstrate that HSF1 can be activated in the absence of physiological stress and also provide support for a model of regulation of HSF1 and HSF2 activity by a titratable negative regulatory factor.

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Temperature-dependent increase in the DNA-binding activity of a heat shock factor in an extract of tobacco cultured cells

Plant Molecular Biology ◽

10.1007/bf00020602 ◽

1996 ◽

Vol 31 (1) ◽

pp. 13-22 ◽

Cited By ~ 6

Author(s):

Sae Shimizu ◽

Yoshinobu Itoh ◽

Ken-ichi Yamazaki

Keyword(s):

Heat Shock ◽

Dna Binding ◽

Cultured Cells ◽

Heat Shock Factor ◽

Binding Activity ◽

Temperature Dependent ◽

Dna Binding Activity

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Uncoupling Stress-Inducible Phosphorylation of Heat Shock Factor 1 from Its Activation

Molecular and Cellular Biology ◽

10.1128/mcb.00816-14 ◽

2015 ◽

Vol 35 (14) ◽

pp. 2530-2540 ◽

Cited By ~ 45

Author(s):

Marek A. Budzyński ◽

Mikael C. Puustinen ◽

Jenny Joutsen ◽

Lea Sistonen

Keyword(s):

Heat Shock ◽

Dna Binding ◽

Posttranslational Modifications ◽

Binding Activity ◽

Heat Shock Factor 1 ◽

Dna Binding Activity ◽

Expression Of Genes ◽

Genes Encoding ◽

Multistep Process ◽

Shock Proteins

In mammals the stress-inducible expression of genes encoding heat shock proteins is under the control of the heat shock transcription factor 1 (HSF1). Activation of HSF1 is a multistep process, involving trimerization, acquisition of DNA-binding and transcriptional activities, which coincide with several posttranslational modifications. Stress-inducible phosphorylation of HSF1, or hyperphosphorylation, which occurs mainly within the regulatory domain (RD), has been proposed as a requirement for HSF-driven transcription and is widely used for assessing HSF1 activation. Nonetheless, the contribution of hyperphosphorylation to the activity of HSF1 remains unknown. In this study, we generated a phosphorylation-deficient HSF1 mutant (HSF1Δ∼PRD), where the 15 known phosphorylation sites within the RD were disrupted. Our results show that the phosphorylation status of the RD does not affect the subcellular localization and DNA-binding activity of HSF1. Surprisingly, under stress conditions, HSF1Δ∼PRD is a potent transactivator of both endogenous targets and a reporter gene, and HSF1Δ∼PRD has a reduced activation threshold. Our results provide the first direct evidence for uncoupling stress-inducible phosphorylation of HSF1 from its activation, and we propose that the phosphorylation signature alone is not an appropriate marker for HSF1 activity.

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