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Heat shock factors (HSFs) are the main transcriptional regulators of the evolutionarily conserved heat shock response. Beyond cell stress, several studies have demonstrated that HSFs also contribute to a vast variety of human pathologies, ranging from metabolic diseases to cancer and neurodegeneration. Despite their evident role in mitigating cellular perturbations, the functions of HSF1 and HSF2 in physiological proteostasis have remained inconclusive. Here, we analyzed a comprehensive selection of paraffin-embedded human tissue samples with immunohistochemistry. We demonstrate that both HSF1 and HSF2 display distinct expression and subcellular localization patterns in benign tissues. HSF1 localizes to the nucleus in all epithelial cell types, whereas nuclear expression of HSF2 was limited to only a few cell types, especially the spermatogonia and the urothelial umbrella cells. We observed a consistent and robust cytoplasmic expression of HSF2 across all studied smooth muscle and endothelial cells, including the smooth muscle cells surrounding the vasculature and the high endothelial venules in lymph nodes. Outstandingly, HSF2 localized specifically at cell-cell adhesion sites in a broad selection of tissue types, such as the cardiac muscle, liver, and epididymis. To the best of our knowledge, this is the first study to systematically describe the expression and localization patterns of HSF1 and HSF2 in benign human tissues. Thus, our work expands the biological landscape of these factors and creates the foundation for the identification of specific roles of HSF1 and HSF2 in normal physiological processes., Competing Interests: Declarations of interest The authors declare no competing interests., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

Defects in cellular protein homeostasis are associated with many severe and prevalent pathological conditions such as neurodegenerative diseases, muscle dystrophies, and metabolic disorders. One way to counteract these defects is to improve the protein homeostasis capacity through induction of the heat shock response. Despite numerous attempts to develop strategies for chemical activation of the heat shock response by heat shock transcription factor 1 (HSF1), the underlying mechanisms of drug candidates' mode of action are poorly understood. To lower the threshold for the heat shock response activation, we used the chaperone co-inducer BGP-15 that was previously shown to have beneficial effects on several proteinopathic disease models. We found that BGP-15 treatment combined with heat stress caused a substantial increase in HSF1-dependent heat shock protein 70 (HSPA1A/B) expression already at a febrile range of temperatures. Moreover, BGP-15 alone inhibited the activity of histone deacetylases (HDACs), thereby increasing chromatin accessibility at multiple genomic loci including the stress-inducible HSPA1A. Intriguingly, treatment with well-known potent HDAC inhibitors trichostatin A and valproic acid enhanced the heat shock response and improved cytoprotection. These results present a new pharmacological strategy for restoring protein homeostasis by inhibiting HDACs, increasing chromatin accessibility, and lowering the threshold for heat shock response activation.

Engagement of death receptors induces caspase activation and apoptosis. A recent study reported altered protein expression, including increased Hsp70 levels during CD95-mediated apoptosis. Here, we examined the mechanism underlying increased Hsp70 levels in cells challenged with a monoclonal antibody directed against the CD95 receptor. Levels of Hsp70 were found to increase in a dose-dependent manner, occurring independently of either heat shock factor 1 activation or the accumulation of Hsp70 messenger ribonucleic acid (mRNA), suggesting the involvement of posttranslational modifications. Inhibition of translation and de novo protein synthesis by cycloheximide resulted in Hsp70 protein levels diminishing over time in control cells, whereas its level remained constant during CD95 signaling. In addition, death receptor activation through exposure of cells to tumor necrosis factor-related apoptosis-inducing ligand did not alter Hsp70 levels. These findings demonstrate that receptor-specific signaling through the CD95 increases the stability of Hsp70 protein, rather than mRNA, when compared with control cells. The results describe a novel mechanism of heat shock protein accumulation, where increased protein stability and reduced turnover, is the mechanism by which Hsp70 accumulates in cells during CD95-mediated apoptosis.