Comprehensive analysis of human tissues reveals unique expression and localization patterns of HSF1 and HSF2.

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.

Stress biology: Complexity and multifariousness in health and disease.

Preserving and regulating cellular homeostasis in the light of changing environmental conditions or developmental processes is of pivotal importance for single cellular and multicellular organisms alike. To counteract an imbalance in cellular homeostasis transcriptional programs evolved, called the heat shock response, unfolded protein response, and integrated stress response, that act cell-autonomously in most cells but in multicellular organisms are subjected to cell-nonautonomous regulation. These transcriptional programs downregulate the expression of most genes but increase the expression of heat shock genes, including genes encoding molecular chaperones and proteases, proteins involved in the repair of stress-induced damage to macromolecules and cellular structures. Sixty-one years after the discovery of the heat shock response by Ferruccio Ritossa, many aspects of stress biology are still enigmatic. Recent progress in the understanding of stress responses and molecular chaperones was reported at the 12th International Symposium on Heat Shock Proteins in Biology, Medicine and the Environment in the Old Town Alexandria, VA, USA from 28th to 31st of October 2023.

Transcriptional reprogramming at the intersection of the heat shock response and proteostasis.

Cellular homeostasis is constantly challenged by a myriad of extrinsic and intrinsic stressors. To mitigate the stress-induced damage, cells activate transient survival programs. The heat shock response (HSR) is an evolutionarily well-conserved survival program that is activated in response to proteotoxic stress. The HSR encompasses a dual regulation of transcription, characterized by rapid activation of genes encoding molecular chaperones and concomitant global attenuation of non-chaperone genes. Recent genome-wide approaches have delineated the molecular depth of stress-induced transcriptional reprogramming. The dramatic rewiring of gene and enhancer networks is driven by key transcription factors, including heat shock factors (HSFs), that together with chromatin-modifying enzymes remodel the 3D chromatin architecture, determining the selection of either gene activation or repression. Here, we highlight the current advancements of molecular mechanisms driving transcriptional reprogramming during acute heat stress. We also discuss the emerging implications of HSF-mediated stress signaling in the context of physiological and pathological conditions.

A rare and complete response to combination therapy with radiation and nivolumab in a patient with metastatic urothelial cancer.

According to the current understanding, radiotherapy can enhance the effectiveness of cancer immunotherapy due to radiation-induced release of tumour-associated antigens. Here, we present a case with a metastatic urothelial carcinoma who received nivolumab and palliative radiotherapy to a residual tumour in the vagina and to a large metastatic visceral lymph node. The treatment resulted in a rapid and virtually complete response for the time being in all metastases and in the large parailiac tumour mass. Follow up continues. The presented case demonstrates that the combinatory treatment with radiotherapy and immunotherapy can result in an exceptional response for the benefit of the patient with urothelial cancer. To our knowledge, this is one of the largest metastatic masses to disappear with a combination of immuno-oncologic (nivolumab) and radiation therapies.

Heat Shock Factor 2 Protects against Proteotoxicity by Maintaining Cell-Cell Adhesion.

Maintenance of protein homeostasis, through inducible expression of molecular chaperones, is essential for cell survival under protein-damaging conditions. The expression and DNA-binding activity of heat shock factor 2 (HSF2), a member of the heat shock transcription factor family, increase upon exposure to prolonged proteotoxicity. Nevertheless, the specific roles of HSF2 and the global HSF2-dependent gene expression profile during sustained stress have remained unknown. Here, we found that HSF2 is critical for cell survival during prolonged proteotoxicity. Strikingly, our RNA sequencing (RNA-seq) analyses revealed that impaired viability of HSF2-deficient cells is not caused by inadequate induction of molecular chaperones but is due to marked downregulation of cadherin superfamily genes. We demonstrate that HSF2-dependent maintenance of cadherin-mediated cell-cell adhesion is required for protection against stress induced by proteasome inhibition. This study identifies HSF2 as a key regulator of cadherin superfamily genes and defines cell-cell adhesion as a determinant of proteotoxic stress resistance.

Tailoring of Proteostasis Networks with Heat Shock Factors.

Heat shock factors (HSFs) are the main transcriptional regulators of the heat shock response and indispensable for maintaining cellular proteostasis. HSFs mediate their protective functions through diverse genetic programs, which are composed of genes encoding molecular chaperones and other genes crucial for cell survival. The mechanisms that are used to tailor HSF-driven proteostasis networks are not yet completely understood, but they likely comprise from distinct combinations of both genetic and proteomic determinants. In this review, we highlight the versatile HSF-mediated cellular functions that extend from cellular stress responses to various physiological and pathological processes, and we underline the key advancements that have been achieved in the field of HSF research during the last decade.

Global SUMOylation on active chromatin is an acute heat stress response restricting transcription.

BACKGROUND: Cells have developed many ways to cope with external stress. One distinctive feature in acute proteotoxic stresses, such as heat shock (HS), is rapid post-translational modification of proteins by SUMOs (small ubiquitin-like modifier proteins; SUMOylation). While many of the SUMO targets are chromatin proteins, there is scarce information on chromatin binding of SUMOylated proteins in HS and the role of chromatin SUMOylation in the regulation of transcription. RESULTS: We mapped HS-induced genome-wide changes in chromatin occupancy of SUMO-2/3-modified proteins in K562 and VCaP cells using ChIP-seq. Chromatin SUMOylation was further correlated with HS-induced global changes in transcription using GRO-seq and RNA polymerase II (Pol2) ChIP-seq along with ENCODE data for K562 cells. HS induced a rapid and massive rearrangement of chromatin SUMOylation pattern: SUMOylation was gained at active promoters and enhancers associated with multiple transcription factors, including heat shock factor 1. Concomitant loss of SUMOylation occurred at inactive intergenic chromatin regions that were associated with CTCF-cohesin complex and SETDB1 methyltransferase complex. In addition, HS triggered a dynamic chromatin binding of SUMO ligase PIAS1, especially onto promoters. The HS-induced SUMOylation on chromatin was most notable at promoters of transcribed genes where it positively correlated with active transcription and Pol2 promoter-proximal pausing. Furthermore, silencing of SUMOylation machinery either by depletion of UBC9 or PIAS1 enhanced expression of HS-induced genes. CONCLUSIONS: HS-triggered SUMOylation targets promoters and enhancers of actively transcribed genes where it restricts the transcriptional activity of the HS-induced genes. PIAS1-mediated promoter SUMOylation is likely to regulate Pol2-associated factors in HS.

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

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.