Activation Of Heat Shock Factor Research Articles

N-butyrate increases heat shock protein 70 through heat shock factor 1 and AMP-activated protein kinase pathways in human intestinal Caco-2 cells

Impaired integrity of the intestinal epithelium is a cause of intestinal and extraintestinal diseases. Heat shock protein 70 (HSP70), a cytoprotective protein, plays an important role in maintaining intestinal homeostasis. The intestinal expression of HSP70 is linked with the local microbiota. The present study investigated the molecular mechanisms underlying the upregulation of HSP70 by n-butyrate, a major metabolite of the intestinal microbiota in human intestinal Caco-2 cells. Treatment of Caco-2 cells with n-butyrate upregulated HSP70 protein and mRNA levels in a dose-dependent manner. Using luciferase reporter assay, it was found that n-butyrate enhanced the transcriptional activity of HSP70. These effects were sensitive to the inhibition of heat shock factor 1 (HSF1), a transcription factor, and AMP-activated protein kinase (AMPK). N-butyrate increased the phosphorylation (activity) of HSF1 and AMPK. Taken together, this study shows that n-butyrate is partly involved in the microbiota-dependent intestinal expression of HSP70, and the effect is exerted through the HSF1 and AMPK pathways.

L-Theanine attenuates heat stress-induced proteotoxicity and alterations in carbohydrate and lipid metabolism via heat shock factor 1.

Extreme heat caused by global warming accelerated the frequency of heat stress (HS). Proteotoxic stress induced by the aggregation of misfolded proteins and metabolic stress triggered by alterations in the metabolism were observed during HS. The activation of heat shock factor 1 (Hsf1) and its interaction with adenosine 5'-monophosphate (AMP)-activated protein kinase (AMPK) are critical in addressing proteotoxicity and metabolic stress in heat-stressed organisms. Previous studies have shown that L-theanine (LTA) can regulate nutrient metabolism through the AMPK pathway and can alleviate HS. Therefore, we hypothesize that LTA may help in restoring homeostasis by regulating nutrient metabolism under HS. Here, we investigated the effects of LTA on nutrient metabolism in heat-stressed rats and characterized the underlying mechanisms using RNA sequencing and metabonomics. The results showed that LTA alleviated HS-induced liver damage, promoted body weight gain, decreased serum cortisol and enhanced the total protein content. Besides, it regulated the expression of genes related to carbohydrate, lipid and amino acid metabolism and altered metabolite levels. Moreover, LTA inhibited the expression of Hsf1 and heat shock protein 70 (Hsp70), promoted AMPK phosphorylation and the expression of glucose-6-phosphatase catalytic subunit 1 (G6pc), and inhibited the phosphorylation of acetyl-CoA carboxylase 1 (ACC1) in heat-stressed rats. Mechanistically, LTA alleviated HS-induced proteotoxic stress by acting on Hsf1/Hsp70; simultaneously, it promoted AMPK phosphorylation by suppressing Hsf1 expression, which in turn inhibited fatty acid synthesis and hepatic gluconeogenesis, thus alleviating HS-induced metabolic stress. These results suggest that LTA regulates nutrient metabolism through Hsf1/AMPK and alleviates HS-induced proteotoxicity via Hsf1/Hsp70.

Crosstalk between heat shock factor 1 and signal transducer and activator of transcription 3 mediated by interleukin-8 autocrine signaling maintains the cancer stem cell phenotype in liver cancer.

Liver cancer stem cells (LCSCs) cause therapeutic refractoriness and relapse in hepatocellular carcinoma. Heat shock factor 1 (HSF1) plays versatile roles in multiple cancers. However, the role of HSF1 in LCSCs is not well understood. This study investigated the function and signal mechanisms of HSF1 in maintaining LCSC phenotypes. We established two LCSC lines, HepG2-R and HuH-7-R. Constitutive activation of HSF1 was observed in these LCSCs. Specific short hairpin RNAs (shRNAs) and chemical inhibitors were used to identify the relationship between HSF1 expression and LCSCs phenotypes. We revealed a concomitant activation modality involving HSF1 and STAT3 in LCSCs and liver cancer tissues. We also found that liver cancer patients whose HSF1 and STAT3 mRNA expression levels were high presented with unfavorable clinicopathological characteristics. Moreover, the secretion of interleukin-8 (IL-8) was elevated in the LCSC medium and was directly regulated by HSF1 at the transcriptional level. In turn, IL-8 activated HSF1 and STAT3 signaling, and a neutralizing IL-8 antibody inhibited HSF1 and STAT3 activity, reduced cancer stem cell marker expression, and decreased LCSC microsphere formation. Simultaneous intervention with HSF1 and STAT3 led to synergistically suppressed stemness acquisition and growth suppression in the LCSCs in vivo and in vitro. Our study indicates that IL-8 mediates the crosstalk between the HSF1 and Stat3 signaling pathways in LCSCs and that the combined targeting of HSF1 and STAT3 is a promising treatment strategy for patients with advanced liver cancer.

Heat shock factor HSF1 regulates BDNF gene promoters upon acute stress in the hippocampus, together with pCREB.

Heat shock factor 1 (HSF1) is a master stress-responsive transcriptional factor, protecting cells from death. However, its gene regulation in vivo in the brain in response to neuronal stimuli remains elusive. Here, we investigated its direct regulation of the brain-derived neurotrophic factor (BDNF) gene (Bdnf) in response to acute neuronal stress stimuli in the brain. The results of immunohistochemistry and chromatin immunoprecipitation quantitative PCR (ChIP-qPCR) showed that administration of kainic acid (a glutamate receptor agonist inducing excitotoxity) to young adult mice induced HSF1 nuclear translocation and its binding to multiple Bdnf promoters in the hippocampus. Footshock, a physical stressor used for learning, also induced HSF1 binding to selected Bdnf promoters I and IV. This is, to our knowledge, the first demonstration of HSF1 gene regulation in response to neuronal stimuli in the hippocampus in vivo. HSF1 binding sites (HSEs) in Bdnf promoters I and IV were also detected when immunoprecipitated by an antibody of phosphorylated (p)CREB (cAMP-responsive element-binding protein), suggesting their possible interplay in acute stress-induced Bdnf transcription. Interestingly, their promoter binding patterns differed by KA and footshock, suggesting that HSF1 and pCREB orchestrate to render fine-tuned promoter control depending on the types of stress. Further, HSF1 overexpression increased Bdnf promoter activity in a luciferase assay, while virus infection of constitutively active-form HSF1 increased levels of BDNF mRNA and protein in vitro in primary cultured neurons. These results indicated that HSF1 activation of Bdnf promoter was sufficient to induce BDNF expression. Taken together, these results suggest that HSF1 promoter-specific control of Bdnf gene regulation plays an important role in neuronal protection and plasticity in the hippocampus in response to acute stress, possibly interplaying with pCREB.

Abstract 15254: Acta2 Missense Variant Increases Atherosclerotic Plaque Burdern by Heat Shock Factor 1-Driven Augmented Phenotypic Modulation of Smooth Muscle Cells

ACTA2 encodes the contractile protein smooth muscle α-actin expressed in smooth muscle cells (SMCs). A heterozygous pathogenic variant in ACTA2 , p.R149C, predisposes to premature coronary artery disease (CAD) in the absence of risk factors. Recent studies have determined that SMCs contribute to atherosclerotic plaque formation after undergoing phenotypic modulation, characterized by de-differentiation and increased expression of macrophage, fibroblast, stem, and osteogenic markers. This SMC modulation is driven in part by exogenous cholesterol entering the endoplasmic reticulum (ER) and activating ER stress, specifically PERK signaling. To determine how a mutation in SM α-actin predisposes to atherosclerosis, Acta2 R149C/+ mice were crossed into Apoe -/- mice and fed a high fat diet for 3 months. Despite no differences in serum lipid levels, the Acta2 R149C/+ Apoe -/- mice have a 3-fold increased plaque burden compared to Apoe -/- mice. SMCs explanted from Acta2 R149C/+ mice have phenotypic modulation at baseline while WT SMCs do not. We determined that misfolding of the R149C SMα-actin activates heat shock factor 1 (HSF1), which increases endogenous cholesterol biosynthesis through increasing HMG-CoA reductase expression. The increase in endogenous cholesterol levels induces ER stress, activates PERK, and augments both modulation of SMCs and atherosclerotic plaque formation. ScRNA sequencing confirms increased HSF1 and PERK signaling and modulation in SMCs in the mutant mice. Importantly, pravastatin treatment dramatically lowers plaque burden to similar low levels of plaque burden in both the Acta2 R149C/+ Apoe -/- and Apoe -/- mice (Fig. 1a) . These studies identify a novel mechanism by which mutant SM α-actin increases endogenous cholesterol biosynthesis and drives atherosclerotic plaque formation, a mechanism that also informs as to why statins are effective in individuals at risk for CAD who do not have elevated plasma cholesterol levels (Fig. 1b) .

The HSF1-CPT1a Pathway Is Differentially Regulated in NAFLD Progression.

Obesity and obesity-associated diseases represent one of the key health challenges of our time. In this context, aberrant hepatic lipid accumulation is a central pathological aspect of non-alcoholic fatty liver disease (NAFLD). By comparing methylation signatures of liver biopsies before and after bariatric surgery, we recently demonstrated the strong enrichment of differentially methylated heat shock factor 1 (HSF1) binding sites (>400-fold) in the process of liver remodeling, indicating a crucial role of HSF1 in modulating central aspects of NAFLD pathogenesis. Using cellular models of NAFLD, we were able to show that HSF1 is activated during fat accumulation in hepatocytes, mimicking conditions in patients before bariatric surgery. This induction was abolished by starving the cells, mimicking the situation after bariatric surgery. Regarding this connection, carnitine palmitoyltransferase 1 isoform A (CTP1a), a central regulator of lipid beta-oxidation, was identified as a HSF1 target gene by promoter analysis and HSF1 knockdown experiments. Finally, pharmacological activation of HSF1 through celastrol reduced fat accumulation in the cells in a HSF1-dependent manner. In conclusion, we were able to confirm the relevance of HSF1 activity and described a functional HSF1-CPT1a pathway in NAFLD pathogenesis.

BRCA mutational status shapes the stromal microenvironment of pancreatic cancer linking clusterin expression in cancer associated fibroblasts with HSF1 signaling

Tumors initiate by mutations in cancer cells, and progress through interactions of the cancer cells with non-malignant cells of the tumor microenvironment. Major players in the tumor microenvironment are cancer-associated fibroblasts (CAFs), which support tumor malignancy, and comprise up to 90% of the tumor mass in pancreatic cancer. CAFs are transcriptionally rewired by cancer cells. Whether this rewiring is differentially affected by different mutations in cancer cells is largely unknown. Here we address this question by dissecting the stromal landscape of BRCA-mutated and BRCA Wild-type pancreatic ductal adenocarcinoma. We comprehensively analyze pancreatic cancer samples from 42 patients, revealing different CAF subtype compositions in germline BRCA-mutated vs. BRCA Wild-type tumors. In particular, we detect an increase in a subset of immune-regulatory clusterin-positive CAFs in BRCA-mutated tumors. Using cancer organoids and mouse models we show that this process is mediated through activation of heat-shock factor 1, the transcriptional regulator of clusterin. Our findings unravel a dimension of stromal heterogeneity influenced by germline mutations in cancer cells, with direct implications for clinical research.

Loss of heat shock factor 1 promotes hepatic stellate cell activation and drives liver fibrosis.

Liver fibrosis is an aberrant wound healing response that results from chronic injury and is mediated by hepatocellular death and activation of hepatic stellate cells (HSCs). While induction of oxidative stress is well established in fibrotic livers, there is limited information on stress‐mediated mechanisms of HSC activation. Cellular stress triggers an adaptive defense mechanism via master protein homeostasis regulator, heat shock factor 1 (HSF1), which induces heat shock proteins to respond to proteotoxic stress. Although the importance of HSF1 in restoring cellular homeostasis is well‐established, its potential role in liver fibrosis is unknown. Here, we show that HSF1 messenger RNA is induced in human cirrhotic and murine fibrotic livers. Hepatocytes exhibit nuclear HSF1, whereas stellate cells expressing alpha smooth muscle actin do not express nuclear HSF1 in human cirrhosis. Interestingly, despite nuclear HSF1, murine fibrotic livers did not show induction of HSF1 DNA binding activity compared with controls. HSF1‐deficient mice exhibit augmented HSC activation and fibrosis despite limited pro‐inflammatory cytokine response and display delayed fibrosis resolution. Stellate cell and hepatocyte‐specific HSF1 knockout mice exhibit higher induction of profibrogenic response, suggesting an important role for HSF1 in HSC activation and fibrosis. Stable expression of dominant negative HSF1 promotes fibrogenic activation of HSCs. Overactivation of HSF1 decreased phosphorylation of JNK and prevented HSC activation, supporting a protective role for HSF1. Our findings identify an unconventional role for HSF1 in liver fibrosis. Conclusion: Our results show that deficiency of HSF1 is associated with exacerbated HSC activation promoting liver fibrosis, whereas activation of HSF1 prevents profibrogenic HSC activation.

Abstract ND08: NXP800: A first-in-class orally active, small-molecule HSF1 pathway inhibitor

Abstract We report the discovery of NXP800 — a first-in-class orally active HSF1 pathway inhibitor identified in our academic laboratory at the Institute of Cancer Research, London — and its progression into Phase I clinical trials in collaboration with our partner Nuvectis Pharma. HSF1 (Heat Shock Factor 1) is an ancient stress-inducible transcription factor that plays a key role in the transcriptional activation of the eukaryotic heat shock response. This leads to expression of many heat shock proteins, including molecular chaperones, and through this HSF1 acts as a master transcriptional regulator of proteostasis. Importantly, HSF1 is hijacked by cancer cells to activate a set of genes that overlaps with, but is not identical to, the classical heat shock response. The HSF1 pathway has been shown to play a key role in oncogenesis and the hallmark features of malignancy and is important in the initiation and progression of many experimental cancer models. Importantly, while HSF1 has been demonstrated to be important in mouse models of oncogenesis, knockout of the homologue of HSF1 has only modest effects in mice and flies under non-stressed laboratory conditions – supporting the potential for a therapeutic window. In addition, gene amplification, expression and activation of HSF1 has been shown to predict clinical outcome in many human cancers. Despite strong scientific rationale and pre-clinical validation, pharmaceutical exploitation has been limited because analysis of the available molecular structures of HSF1 shows it to be very difficult to drug. Because of this, we set out to discover inhibitors of the HSF1 pathway using an initial phenotypic screen that reports on the activation of the HSF1-regulated gene HSP72. This identified the bisamide series that was followed up by a detailed understanding of structure-activity relationships and medicinal chemistry optimization, particularly to improve pharmaceutical properties, leading to the selection of NXP800 (CCT361814) as the development candidate. NXP800 has good pharmacokinetic properties in mice, including oral bioavailability; also, pharmacodynamic studies show modulation of biomarkers of the HSF1 pathway and stress response in human tumour xenograft models. NXP800 exhibits striking therapeutic activity in xenografts of human ovarian clear cell ovarian cancer (OCCC) and endometrioid ovarian cancer, two serious conditions of high unmet medical need with limited treatment options – including prolonged tumour growth inhibition and regression at doses that show good toleration. The efficacy and tolerability data indicate a clear therapeutic window, supported by selectivity and safety profiting in pharmacology and kinome panels. In addition, we identified deficiency in ARID1A, a component of the SWI/SNF chromatin remodelling complex, as indicative of greater therapeutic responsiveness. This was confirmed through broader evaluation in human cancer cell line panels that also indicates therapeutic potential for NXP800 in additional cancer types. Studies to support mechanism of action and identify patient selection biomarkers will be described. A Phase 1 trial of NXP800 is now open, incorporating validated predictive, pharmacokinetic and pharmacodynamic biomarkers that provide a robust Pharmacological Audit Trail and support the prediction of dose-to-human. Citation Format: Paul Workman. NXP800: A first-in-class orally active, small-molecule HSF1 pathway inhibitor [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr ND08.

Hyperthermia increases HSP production in human PDMCs by stimulating ROS formation, p38 MAPK and Akt signaling, and increasing HSF1 activity

BackgroundHuman placenta-derived multipotent cells (hPDMCs) are isolated from a source uncomplicated by ethical issues and are ideal for therapeutic applications because of their capacity for multilineage differentiation and proven immunosuppressive properties. It is known that heat shock preconditioning induces the upregulation of heat shock proteins (HSPs), which enhance survival and engraftment of embryonic stem cells (ESCs) during transplantation in live animal models, although whether heat shock preconditioning has the same effects in hPDMCs is unclear.MethodsThe hPDMCs were isolated from placenta of healthy donors. The cells were treated with heat shock (43 °C, 15 min), followed by evaluation of cell viability. Furthermore, the HSPs expression was assessed by Western blot, qPCR. The reactive oxygen species (ROS) production and signal pathway activation were determined by flow cytometry and Western blot, respectively. The regulatory pathways involved in HSPs expression were examined by pretreatment with chemical inhibitors, and siRNAs of MAPK, Akt, and heat shock factor 1 (HSF1), followed by determination of HSPs expression.ResultsThis study demonstrates that heat shock treatment induced ROS generation and HPSs expression in hPDMCs. Heat shock stimulation also increased p38 MAPK and Akt phosphorylation. These effects were reduced by inhibitors of ROS, p38 MAPK and Akt. Moreover, we found that heat shock treatment enhanced nuclear translocation of the HSF1 in hPDMCs, representing activation of HSF1. Pretreatment of hPDMCs with ROS scavengers, SB203580 and Akt inhibitors also reduced the translocation of HSF1 induced by heat shock.ConclusionsOur data indicate that heat shock acts via ROS to activate p38 MAPK and Akt signaling, which subsequently activates HSF1, leading to HSP activation and contributing to the protective role of hPDMCs.

Molecular Bases of Heat Stress Responses in Vegetable Crops With Focusing on Heat Shock Factors and Heat Shock Proteins.

The effects of the climate change including an increase in the average global temperatures, and abnormal weather events such as frequent and severe heatwaves are emerging as a worldwide ecological concern due to their impacts on plant vegetation and crop productivity. In this review, the molecular processes of plants in response to heat stress—from the sensing of heat stress, the subsequent molecular cascades associated with the activation of heat shock factors and their primary targets (heat shock proteins), to the cellular responses—have been summarized with an emphasis on the classification and functions of heat shock proteins. Vegetables contain many essential vitamins, minerals, antioxidants, and fibers that provide many critical health benefits to humans. The adverse effects of heat stress on vegetable growth can be alleviated by developing vegetable crops with enhanced thermotolerance with the aid of various genetic tools. To achieve this goal, a solid understanding of the molecular and/or cellular mechanisms underlying various responses of vegetables to high temperature is imperative. Therefore, efforts to identify heat stress-responsive genes including those that code for heat shock factors and heat shock proteins, their functional roles in vegetable crops, and also their application to developing vegetables tolerant to heat stress are discussed.

AKT1 mediates multiple phosphorylation events that functionally promote HSF1 activation.

The heat stress response activates the transcription factor heat shock factor 1 (HSF1), which subsequently upregulates heat shock proteins to maintain the integrity of the proteome. HSF1 activation requires nuclear localization, trimerization, DNA binding, phosphorylation and gene transactivation. Phosphorylation at S326 is an important regulator of HSF1 transcriptional activity. Phosphorylation at S326 is mediated by AKT1, mTOR, p38, MEK1 and DYRK2. Here, we observed activation of HSF1 by AKT1 independently of mTOR. AKT2 also phosphorylated S326 of HSF1 but showed weak ability to activate HSF1. Similarly, mTOR, p38, MEK1 and DYRK2 all phosphorylated S326 but AKT1 was the most potent activator. Mass spectrometry showed that AKT1 also phosphorylated HSF1 at T142, S230 and T527 in addition to S326, whereas the other kinases did not. Subsequent investigation revealed that phosphorylation at T142 is necessary for HSF1 trimerization and that S230, S326 and T527 are required for HSF1 gene transactivation and recruitment of TFIIB and CDK9. Interestingly, T527 as a phosphorylated residue has not been previously shown and sits in the transactivation domain, further implying a role for this site in HSF1 gene transactivation. This study suggests that HSF1 hyperphosphorylation is targeted and these specific residues have direct function in regulating HSF1 transcriptional activity.

Heat shock factor 1 inhibition sensitizes pancreatic cancer to gemcitabine via the suppression of cancer stem cell-like properties

Pancreatic cancer is a fatal disease with poor prognosis. Gemcitabine has been regarded as the mainstay of chemotherapy for pancreatic cancer; however, it is accompanied with a high rate of chemoresistance. Cancer stem cells (CSCs) are characterized by resistance to traditional chemo- and radiotherapies. We have previously reported that heat shock factor 1 (HSF1) is involved in the invasion and metastasis of pancreatic cancer, a highly conserved transcriptional factor that mediates the canonical proteotoxic stress response. Here, we investigate whether HSF1 contributes to the chemoresistance of pancreatic cancer cells caused by gemcitabine and explore the underlying mechanisms. Genetically engineered mice (LSL-KrasG12D/+; Trp53fl/+; Pdx1-Cre mice), which spontaneously develop pancreatic cancer, were used to examine the sensitivity of pancreatic cancer to gemcitabine in vivo. We found that HSF1 was enriched in sphere-forming cancer cells. Panc-1 and MiaPaCa-2 cells treated chronically with gemcitabine displayed increased transcription and expression of CSC-associated markers. In addition, gemcitabine-surviving Panc-1 and MiaPaCa-2 cells showed an increased ability to form tumorspheres. Moreover, we observed that gemcitabine treatment increased the activity and expression of HSF1, as well as transcription of its downstream targets. Finally, HSF1 inhibition significantly suppressed the expression of CSC-associated markers, augmented the cancer-killing property of gemcitabine, and increased chemosensitivity to gemcitabine in vivo. Our study reveals a novel mechanism in which HSF1 promotes the chemoresistance of pancreatic cancer to gemcitabine by modulating CSC-like properties. Targeting HSF1 could be thus a rational strategy to improve treatment outcomes.

A novel HSF1 activator ameliorates non-alcoholic steatohepatitis by stimulating mitochondrial adaptive oxidation.

Non-alcoholic steatohepatitis (NASH) is the more severe form of metabolic associated fatty liver disease (MAFLD) and no pharmacological treatment as yet been approved. Identification of novel therapeutic targets and their agents is critical to overcome the current inadequacy of drug treatment for NASH. The correlation between heat shock factor 1 (HSF1) levels and the development of NASH and the target genes of HSF1 in hepatocyte were investigated by chromatin-immunoprecipitation sequencing. The effects and mechanisms of SYSU-3d in alleviating NASH were examined in relevant cell models and mouse models (the Ob/Ob mice, high-fat and high-cholesterol diet and the methionine-choline deficient diet-fed mice). The actions of SYSU-3d in vivo were evaluated. HSF1 is progressively reduced with mitochondrial dysfunction in NASH pathogenesis and activation of this transcription factor by its newly identified activator SYSU-3d effectively inhibited all manifestations of NASH in mice. When activated, the phosphorylated HSF1 (Ser326) translocated to nucleus and bound to the promoter of PPARγ coactivator-1α (PGC-1α) to induce mitochondrial biogenesis. Thus, increasing mitochondrial adaptive oxidation and inhibiting oxidative stress. The deletion of HSF1 and PGC-1α or recovery of HSF1 in HSF1-deficiency cells showed the HSF1/PGC-1α pathway was mainly responsible for the anti-NASH effects of SYSU-3d independent of AMP-activated protein kinase (AMPK). Activation of HSF1 is a practical therapeutic approach for NASH treatment via the HSF1/PGC-1α/mitochondrial pathway and SYSU-3d can be considered as a potential candidate for the treatment of NASH.

Salvianolic Acid Ameliorates Pressure Overload-Induced Cardiac Endothelial Dysfunction via Activating HIF1[Formula: see text]/HSF1/CD31 Pathway.

Pressure overload is a major risk factor for various cardiovascular diseases. Disorders of the endothelium are involved in the pathological mechanisms of pressure, and maintaining endothelial function is a practical strategy to alleviate pressure overload-induced cardiac injury. In this study, we provided evidence that salvianolic acid, the active component of Danshen, a traditional Chinese herb medicine, preserved pressure overload-induced cardiac dysfunction via protecting endothelium. Male C57BL/6J mice were imposed with transverse aortic constriction to mimic pressure overload and treated with salvianolic acid (200[Formula: see text]mg/kg/day) or vehicle for 6 weeks. The hemodynamic and cardiac functional parameters were detected by the cardiac catheter and transthoracic echocardiography. The pathological measurements were conducted by heart hematoxylin-eosin, wheat germ agglutinin staining, Masson's trichrome staining, and immunofluorescence staining. Endothelial cell (EC) proliferation was estimated using the Cell Counting Kit-8, EC migration was evaluated by scratched assay, and EC integrity was observed by electron microscope. Salvianolic acid notably inhibited cardiac chamber enlargement, restrained cardiac contractile dysfunction, and repressed cardiac fibrosis caused by chronic pressure overload. Salvianolic acid maintained endothelial tight junction integrity by boosting the expression of CD31. Furthermore, the endothelial protective effect of salvianolic acid against pressure overload is dependent on the activation of hypoxia-inducible factor 1[Formula: see text], which consequently activated heat shock factor 1 and promoted CD31 expression. Our study uncovered that salvianolic acid protected cardiac ECs against pressure overload via a HIF1[Formula: see text]/HSF1/CD31 pathway, indicating a potential appliance of salvianolic acid in hypertensive heart disease.

FTO promotes multiple myeloma progression by posttranscriptional activation of HSF1 in an m6A-YTHDF2-dependent manner

N6-methyladenosine (m6A), as the most pervasive internal modification of eukaryotic mRNA, plays a crucial role in various cancers, but its role in multiple myeloma (MM) pathogenesis has not yet been investigated. In this study, we revealed significantly decreased m6A methylation in plasma cells (PCs) from MM patients and showed that the abnormal m6A level resulted mainly from upregulation of the demethylase fat mass and obesity-associated protein (FTO). Gain- and loss-of-function studies demonstrated that FTO plays a tumor-promoting and pro-metastatic role in MM. Combined m6A and RNA sequencing (RNA-seq) and subsequent validation and functional studies identified heat shock factor 1 (HSF1) as a functional target of FTO-mediated m6A modification. FTO significantly promotes MM cell proliferation, migration, and invasion by targeting HSF1/HSPs in a YTHDF2-dependent manner. FTO inhibition, especially when combined with bortezomib (BTZ) treatment, synergistically inhibited myeloma bone tumor formation and extramedullary spread in NOD-Prkdcem26Cd52il2rgem26Cd22/Nju (NCG) mice. We demonstrated the functional importance of m6A demethylase FTO in MM progression, especially in promoting extramedullary myeloma (EMM) formation, and proposed the FTO-HSF1/HSP axis as a potential novel therapeutic target in MM.

PLOD1 acts as a tumor promoter in glioma via activation of the HSF1 signaling pathway.

Procollagen-lysine, 2-oxoglutarate 5-dioxygenase 1 (PLOD1) is a collagen-related lysyl hydroxylase and its prognostic value in glioma patients was verified. However, its biological function in glioma has yet to be fully investigated. The PLOD1 mRNA status and clinical significance in gliomas were assessed via the GEPIA database. Overexpression or targeted depletion of PLOD1 was carried out in the human glioma cell line U87 and verified by western blotting. CCK8 and colony formation assays were implemented to examine the impact of PLOD1 on the proliferative and colony-forming phenotypes of U87 cells. Luciferase reporter assays and HSF1-specific pharmacologic inhibitors (KRIBB11) were employed to determine the regulatory relationship between PLOD1 and heat shock factor 1 (HSF1). High expression of PLOD1 was observed in tissue samples of glioblastoma multiforme (GBM) and brain lower-grade glioma (LGG). GEPIA overall survival further demonstrated that both GBM and LGG patients with high PLOD1 displayed worse clinical outcomes compared with those with low PLOD1. Overexpression and targeted depletion of PLOD1 enhanced and suppressed U87 cell proliferation and colony formation, respectively. Luciferase reporter assays showed that PLOD1 significantly enhanced the transcriptional activity of HSF1 in HEK293T cells. PLOD1 deficiency in U87 cells inhibited HSF1-induced survivin accumulation, whereas KRIBB11 also blocked the PLOD1-overexpressing induced survivin expression. An inhibitor of HSF1 signaling events abolishedthe increased clonogenic potential caused by PLOD1 overexpression in U87 cells. High expression of PLOD1 can increase the proliferation and colony formation of U87 cells by activating the HSF1 signaling pathway. This study suggested PLOD1/HSF1 as an effective therapeutic target for gliomas.

Heat shock factor 1 in brain tumors: a link with transient receptor potential channels TRPV1 and TRPA1.

Novel data report a "cross-talk" between Heat-Shock Factor 1 (HSF1) and the transient receptor potential vanilloid 1 cation channel (TRPV1) located in the cell membrane, introducing these channels as possible drug targets for the regulation of HSF1 activation. This study aims to investigate the co-expression of TRPV1 and HSF1 in human brain tumors. Additionally, the expression of the transient receptor potential ankyrin 1 channel (TRPA1), which is co-operated with TRPV1 in a plethora of cells, was studied. Immunohistochemical staining for HSF1, TRPV1 and TRPA1 expression was quantitatively analyzed in paraffin-embedded semi-serial tissue sections from 74 gliomas and 71 meningiomas. mRNA levels of HSF1, TRPV1 and TRPA1 were evaluated using real-time PCR. Although HSF1 was significantly increased compared with TRPV1/TRPA1 (p ≤ 0.001) in both gliomas and meningiomas, high co-expression levels for HSF1, TRPV1 and TRPA1 were found in 62.50% of diffuse fibrillary astrocytomas (WHO, grade II), 37.50% of anaplastic astrocytomas (WHO, grade III), 16.32% of glioblastomas multiforme (WHO, grade IV), and 42.25% of meningiomas (WHO, grade I and II). Correlation analysis revealed a relationship of HSF1 with TRPV1/TRPA1 in diffuse fibrillary astrocytomas (WHO, grade II) and benign meningiomas (WHO, grade I) contrary to glioblastomas multiforme (WHO, grade IV) and high grade meningiomas (WHO, grade II). Importantly, TRPA1 and TRPV1 expression levels were significantly increased in meningiomas compared with astrocytic tumors (p < 0.05). In conclusion, HSF1 and TRPV1/TRPA1 co-expression may be implicated in the pathogenesis of human brain tumors and should be considered for the therapeutic approaches for these tumors.

Mechanisms of elevated CO2-induced thermotolerance in plants: the role of phytohormones.

Rising atmospheric CO2 is a key driver of climate change, intensifying drastic changes in meteorological parameters. Plants can sense and respond to changes in environmental parameters including atmospheric CO2 and temperatures. High temperatures beyond the physiological threshold can significantly affect plant growth and development and thus attenuate crop productivity. However, elevated atmospheric CO2 can mitigate the deleterious effects of heat stress on plants. Despite a large body of literature supporting the positive impact of elevated CO2 on thermotolerance, the underlying biological mechanisms and precise molecular pathways that lead to enhanced tolerance to heat stress remain largely unclear. Under heat stress, elevated CO2-induced expression of respiratory burst oxidase homologs (RBOHs) and reactive oxygen species (ROS) signaling play a critical role in stomatal movement, which optimizes gas exchange to enhance photosynthesis and water use efficiency. Notably, elevated CO2 also fortifies antioxidant defense and redox homeostasis to alleviate heat-induced oxidative damage. Both hormone-dependent and independent pathways have been shown to mediate high CO2-induced thermotolerance. The activation of heat-shock factors and subsequent expression of heat-shock proteins are thought to be the essential mechanism downstream of hormone and ROS signaling. Here we review the role of phytohormones in plant response to high atmospheric CO2 and temperatures. We also discuss the potential mechanisms of elevated CO2-induced thermotolerance by focusing on several key phytohormones such as ethylene. Finally, we address some limitations of our current understanding and the need for further research to unveil the yet-unknown crosstalk between plant hormones in mediating high CO2-induced thermotolerance in plants.

Abstract LB204: HSF1 promotes inflammation induced tumor development through ECM remodeling

Abstract HSF1 promotes inflammation induced tumor development through ECM remodelingAbstractIn the colon, long-term exposure to chronic inflammation drives colitis associated colon cancer (CAC) in patients with inflammatory bowel disease (IBD). Chronic inflammation underlies tumor initiation, promotion, invasion, and metastasis. While the causal and clinical link between chronic inflammation and CAC is well established, we lack a molecular understanding of what is the way in which chronic inflammation leads to develop colon cancer. Within the tumor, cancer cells are surrounded by a variety of non-malignant cells, such as macrophages, endothelial cells, neutrophils, cancer-associated fibroblasts (CAFs), and together with the extracellular matrix (ECM) they compose the tumor microenvironment (TME), also termed the stroma. Even the most aggressive cancers depend and interact with their environment mostly through secreted factors. Unlike cancer cells, stromal cells are genomically stable, and do not harbor oncogenic mutations that could drive their co-evolution and functional reprogramming. Rather, stromal reprogramming is thought to be achieved by transcriptional rewiring. Previous work by us and others has shown that the master regulator heat shock factor 1 (HSF1) plays a crucial role in this process, by mediating a transcriptional program in fibroblasts that enables their reprogramming into cancer-associated fibroblasts (CAFs) to promote malignancy. We hypothesizde that HSF1 plays a crucial role in inflammation-driven cancer by initiation of a transcriptional program that leads to changes in the extracellular matrix (ECM). We found that, in cell culture, cancer-induced ECM assembly by fibroblasts requires HSF1. Using an inflammation-driven cancer model in mice, we measured the changes in proteomic and ECM organization over time. We found that HSF1 drives a transcriptional program that leads to ECM remodeling in early stages and results in development of colon cancer. Loss of HSF1 prevents inflammation-induced ECM remodeling. Further to that, in CAC patients, we found high activation of stromal HSF1 and similarity to our HSF1 proteomic ECM signature in human colorectal cancer driven by HSF1. Thus, HSF1-dependent ECM remodeling mediates the transition from chronic inflammation to colon cancer. Citation Format: Oshrat Levi Galibov, Hagar Lavon, Rina Wassermann-Dozorets, Meirav Pevsner-Fischer, Shimrit Mayer, Esther Wershof, Yaniv Stein, Lauren E. Brown, Wenhan Zhang, Gil Friedman, Reinat Nevo, Ofra Golani, Lior H. Katz, Rona Yaeger, Ido Laish, John A. Porco, Erik Sahai, Dror S Shouval, David Kelsen, Ruth Scherz-Shouval. HSF1 promotes inflammation induced tumor development through ECM remodeling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr LB204.