Two Distinct Disulfide Bonds Formed in Human Heat Shock Transcription Factor 1 Act in Opposition To Regulate Its DNA Binding Activity

Abstract

Under circumstances of heat stress, heat shock transcription factor 1 (HSF1) plays important roles in heat shock protein expression. In this study, an increasing concentration of dithiothreitol (DTT) was found to either enhance or inhibit the heat-induced trimerization of HSF1, suggesting the involvement of dual redox-dependent HSF1 activation mechanisms. Our in vitro experiments show that the heat-induced bonding between the cysteine C36 and C103 residues of HSF1 forms an intermolecular disulfide covalent bond (SS-I bond) and that it directly causes HSF1 to trimerize and bond to DNA. Gel filtration assays show that HSF1 can form intermolecular hydrophobic interaction-mediated (iHI-m) noncovalent oligomers. However, the lack of a trimerization domain prevents HSF1 activation, which suggests that iHI-m noncovalent trimerization is a precondition of SS-I bond formation. On the other hand, intramolecular SS-II bond (in which the C153, C373, and C378 residues of HSF1 participate) formation inhibits this iHI-m trimerization, thereby preventing SS-I bond formation and DNA binding. Thus, HSF1 activation is regulated positively by intermolecular SS-I bond formation and negatively by intramolecular SS-II bond formation. Importantly, these two SS bonds confer different DTT sensitivities (the SS-II bond is more sensitive). Therefore, a low concentration of DTT cleaves the SS-II bond but not the SS-I bond and thus improves DNA binding of HSF1, whereas a high concentration DTT cuts both SS bonds and inhibits HSF1 activation. We propose that these interesting effects further explain cellular HSF1 trimerization, DNA binding, and transcription when cells are under stress.

Cellular Stress Responses, The Hormesis Paradigm, and Vitagenes: Novel Targets for Therapeutic Intervention in Neurodegenerative Disorders

“…Thus, an intermolecular disulfide bond formation between C36 and C103 within HSF1 causes trimerization and DNA binding, whereas an intramolecular disulfide bond formation (in which C153, C373, and C378 participate) is inhibitory for the activity of the transcription factor (266).…”

Hormesis, cellular stress response and vitagenes as critical determinants in aging and longevity

“…In addition to thermal stress, the inducible expression of heat shock proteins is also triggered by environmental redox changes or exposure to electrophiles which cause trimerization and DNA binding of HSF1, pointing to the importance of the cysteine redox state for the activation of this transcription factor (Banerjee Mustafi et al, 2009). Thus, an intermolecular disulfide bond formation between C36 and C103 within HSF1 causes trimerization and DNA binding, whereas an intramolecular disulfide bond formation is inhibitory for the activity of the transcription factor (Lu et al, 2008). Hence, pharmacologic modulation of HSF-mediated gene regulation is an emerging area of research which is increasing its potential based on the current knowledge of small-molecule activators and inhibitors of HSFs, so that the impact of HSFs is further extending beyond the heat shock response., this, the potential for attracting growing interest (Akerfelt et al, 2010).…”

Transcription factors Hsf1 and Nrf2 engage in crosstalk for cytoprotection

“…Hsf1 binds DNA as a trimer, whereby each of the DBD of the monomers recognizes the nGAAn sequences in the major groove [80]. Assembly of the DNA binding-competent Hsf1 trimer involves formation of a three-stranded coiled-coil structure, which is generated by intermolecular interactions between the leucine zipper (LZ)1-3 domain [also known as heptad repeat (HR)-A/B region], is negatively regulated by intramolecular interactions between the LZ1-3 and LZ4 (also known as HR-C) domains [81], and is stabilized via intermolecular disulfide bonding between the Hsf1 monomers [64]. The C-terminal transactivation domain (CTAD) is negatively regulated by the regulatory domain (RD).…”