Non-cell-autonomous regulation of germline proteostasis by insulin/IGF-1 signaling-induced dietary peptide uptake via PEPT-1.

Gametogenesis involves active protein synthesis and is proposed to rely on proteostasis. Our previous work in C. elegans indicates that germline development requires coordinated activities of insulin/IGF-1 signaling (IIS) and HSF-1, the central regulator of the heat shock response. However, the downstream mechanisms were not identified. Here, we show that depletion of HSF-1 from germ cells impairs chaperone gene expression, causing protein degradation and aggregation and, consequently, reduced fecundity and gamete quality. Conversely, reduced IIS confers germ cell resilience to HSF-1 depletion-induced protein folding defects and various proteotoxic stresses. Surprisingly, this effect was not mediated by an enhanced stress response, which underlies longevity in low IIS conditions, but by reduced ribosome biogenesis and translation rate. We found that IIS activates the expression of intestinal peptide transporter PEPT-1 by alleviating its repression by FOXO/DAF-16, allowing dietary proteins to be efficiently incorporated into an amino acid pool that fuels germline protein synthesis. Our data suggest this non-cell-autonomous pathway is critical for proteostasis regulation during gametogenesis.

Unveiling the impact of SUMOylation at K298 site of heat shock factor 1 on glioblastoma malignant progression.

BACKGROUND: Glioblastoma (GBM) poses a significant medical challenge due to its aggressive nature and poor prognosis. Mitochondrial unfolded protein response (UPRmt) and the heat shock factor 1 (HSF1) pathway play crucial roles in GBM pathogenesis. Post-translational modifications, such as SUMOylation, regulate the mechanism of action of HSF1 and may influence the progression of GBM. Understanding the interplay between SUMOylation-modified HSF1 and GBM pathophysiology is essential for developing targeted therapies. METHODS: We conducted a comprehensive investigation using cellular, molecular, and in vivo techniques. Cell culture experiments involved establishing stable cell lines, protein extraction, Western blotting, co-immunoprecipitation, and immunofluorescence analysis. Mass spectrometry was utilized for protein interaction studies. Computational modeling techniques were employed for protein structure analysis. Plasmid construction and lentiviral transfection facilitated the manipulation of HSF1 SUMOylation. In vivo studies employed xenograft models for tumor growth assessment. RESULTS: Our research findings indicate that HSF1 primarily undergoes SUMOylation at the lysine residue K298, enhancing its nuclear translocation, stability, and downstream heat shock protein expression, while having no effect on its trimer conformation. SUMOylated HSF1 promoted the UPRmt pathway, leading to increased GBM cell proliferation, migration, invasion, and reduced apoptosis. In vivo studies have confirmed that SUMOylation of HSF1 enhances its oncogenic effect in promoting tumor growth in GBM xenograft models. CONCLUSION: This study elucidates the significance of SUMOylation modification of HSF1 in driving GBM progression. Targeting SUMOylated HSF1 may offer a novel therapeutic approach for GBM treatment. Further investigation into the specific molecular mechanisms influenced by SUMOylated HSF1 is warranted for the development of effective targeted therapies to improve outcomes for GBM patients.

Cadmium causes cerebral mitochondrial dysfunction through regulating mitochondrial HSF1.

Mitochondria, as the powerhouse of the cell, play a vital role in maintaining cellular energy homeostasis and are known to be a primary target of cadmium (Cd) toxicity. The improper targeting of proteins to mitochondria can compromise the normal functions of the mitochondria. However, the precise mechanism by which protein localization contributes to the development of mitochondrial dysfunction induced by Cd is still not fully understood. For this research, Hy-Line white variety chicks (1-day-old) were used and equally distributed into 4 groups: the Control group (fed with a basic diet), the Cd35 group (basic diet with 35 mg/kg CdCl2), the Cd70 group (basic diet with 70 mg/kg CdCl2) and the Cd140 group (basic diet with 140 mg/kg CdCl2), respectively for 90 days. It was found that Cd caused the accumulation of heat shock factor 1 (HSF1) in the mitochondria, and the overexpression of HSF1 in the mitochondria led to mitochondrial dysfunction and neuronal damage. This process is due to the mitochondrial HSF1 (mtHSF1), causing mitochondrial fission through the upregulation of dynamin-related protein 1 (Drp1) content, while inhibiting oligomer formation of single-stranded DNA-binding protein 1 (SSBP1), resulting in the mitochondrial DNA (mtDNA) deletion. The findings unveil an unforeseen role of HSF1 in triggering mitochondrial dysfunction.

Protein Misfolding Releases Human HSF1 from HSP70 Latency Control.

Heat shock factor 1 (HSF1) responds to stress to mount the heat shock response (HSR), a conserved transcriptional program that allows cells to maintain proteostasis by upregulating heat shock proteins (HSPs). The homeostatic stress regulation of HSF1 plays a key role in human physiology and health but its mechanism has remained difficult to pinpoint. Recent work in the budding yeast model has implicated stress-inducible chaperones of the HSP70 family as direct negative regulators of HSF1 activity. Here, we have investigated the latency control and activation of human HSF1 by HSP70 and misfolded proteins. Purified oligomeric HSF1-HSP70 (HSPA1A) complexes exhibited basal DNA binding activity that was inhibited by increasing the levels of HSP70 and, importantly, misfolded proteins reverted the inhibitory effect. Using site-specific UV photo-crosslinking, we monitored HSP70-HSF1 complexes in HEK293T cells. While HSF1 was bound by the substrate binding domain of HSP70 in unstressed cells, activation of HSF1 by heat shock as well as by inducing the misfolding of newly synthesized proteins resulted in release of HSF1 from the chaperone. Taken our results together, we conclude that latent HSF1 populate dynamic complexes with HSP70, which are sensitive to increased levels of misfolded proteins that compete for binding to the HSP70 substrate binding domain. Thus, human HSF1 is activated by various stress conditions that all titrate available HSP70.

Age-dependent heat shock hormesis to HSF-1 deficiency suggests a compensatory mechanism mediated by the unfolded protein response and innate immunity in young Caenorhabditis elegans.

The transcription factor HSF-1 (heat shock factor 1) acts as a master regulator of heat shock response in eukaryotic cells to maintain cellular proteostasis. The protein has a protective role in preventing cells from undergoing ageing, and neurodegeneration, and also mediates tumorigenesis. Thus, modulating HSF-1 activity in humans has a promising therapeutic potential for treating these pathologies. Loss of HSF-1 function is usually associated with impaired stress tolerance. Contrary to this conventional knowledge, we show here that inactivation of HSF-1 in the nematode Caenorhabditis elegans results in increased thermotolerance at young adult stages, whereas HSF-1 deficiency in animals passing early adult stages indeed leads to decreased thermotolerance, as compared to wild-type. Furthermore, a gene expression analysis supports that in young adults, distinct cellular stress response and immunity-related signaling pathways become induced upon HSF-1 deficiency. We also demonstrate that increased tolerance to proteotoxic stress in HSF-1-depleted young worms requires the activity of the unfolded protein response of the endoplasmic reticulum and the SKN-1/Nrf2-mediated oxidative stress response pathway, as well as an innate immunity-related pathway, suggesting a mutual compensatory interaction between HSF-1 and these conserved stress response systems. A similar compensatory molecular network is likely to also operate in higher animal taxa, raising the possibility of an unexpected outcome when HSF-1 activity is manipulated in humans.

HDAC6 modulates the cognitive behavioral function and hippocampal tissue pathological changes of APP/PS1 transgenic mice through HSP90-HSF1 pathway.

The aim of this study was to investigate histone deacetylase 6 (HDAC6) modifies the heat shock protein 90 (HSP90) and heat shock transcription factor 1 (HSF1) affect the levels of pathological markers such as Aß oligomers (Aßo) and Tau phosphorylation (p-Tau) in APP/PS1 double transgenic mice hippocampal tissues or HT22 neurons as well as the changes in cognitive behavioral functions of mice. (1) APP/PS1 transgenic mice (6 months old, 25 ~ 30 g) were randomly assigned to 5 experimental groups, C57BL/6J mice (6 months old, 25 ~ 30 g) were used as 4 control groups, with 8 mice in each group. All mice underwent intracerebroventricular (i.c.v.) cannulation, and the experimental groups were administered with normal saline (APP + NS group), HDAC6 agonist tubastatin A hydrochloride (TSA) (APP + TSA group) or HDAC6 agonist theophylline (Theo) (APP + Theo group), HSP90 inhibitor Ganetespib (Gane) (APP + Gane group), or a combination of pre-injected Gane by TSA (APP + Gane + TSA group); the control group received i.c.v. injections of Gane (Gane group), TSA (TSA group), Theo (Theo group) or NS (NS group), respectively. (2) Mouse hippocampal neurons HT22 were randomly divided into a control group (Control) and an Aß1-42 intervention group (Aß). Within the Aß group, further divisions were made for knockdown HSP90 (Aß + siHSP90 group), overexpression HSP90 (Aß + OE-HSP90 group), knockdown HSF1(Aß + siHSF1 group) and knockdown HSF1 followed by overexpression HSP90 (Aß + siHSF1 + OE-HSP90 group), resulting in a total of 6 groups. Morris water maze test was used to evaluate the cognitive behavior of the mice. Western blot and immunohistochemistry or immunofluorescence were performed to detect the levels of HDAC6, HSP90, HSF1, Aß1-42, Tau protein, and p-Tau in the hippocampal tissue or HT22 cells. qRT-PCR was used to measure the levels of hdac6, hsp90, and hsf1 mRNA in the hippocampus or nerve cells. (1) The levels of HDAC6, Aß1-42 and p-Tau were elevated, while HSP90 and HSF1 were decreased in the hippocampal tissue of APP/PS1 transgenic mice (all P < 0.01). Inhibiting HDAC6 upregulated the expressions of HSP90 and HSF1 in the hippocampal tissue of APP/PS1 mice, while decreasing the levels of Aß1-42 and p-Tau as well as improving the spatial cognitive behavior in mice (P < 0.05 or P < 0.01). The opposite effects were observed upon HDAC6 activation. However, inhibiting HSP90 reduced the expression of HSF1 (P < 0.01) and increased the levels of Aß1-42 and p-Tau (P < 0.05 or P < 0.01) but did not significantly affect the expression of HDAC6 (P > 0.05). No significant changes were observed in the aforementioned indicators in the 4 control groups (P > 0.05). (2) In the Aß1-42 intervention group, HDAC6 and Aß1-42, p-Tau expression levels were elevated, while HSP90 and HSF1 expressions were all decreased, and cell viability was reduced (P < 0.05 or P < 0.01). Overexpression of HSP90 upregulated HSF1 expression, decreased the levels of Aß1-42 and p-Tau, and increased cell viability (P < 0.05 or P < 0.01). Knocking down HSP90 had the opposite effect; and knocking down HSF1 increased the levels of Aß1-42 and p-Tau and decreased cells viability (all P < 0.01), but did not result in significant changes in the expression levels of HSP90 (P > 0.05). Inhibiting HDAC6 can upregulate the expressions of HSP90 and HSF1 but reduce the levels of Aß1-42 and p-Tau in the hippocampus of APP/PS1 mice and improvement of cognitive behavioral function in mice; Overexpression of HSP90 can increase HSF1 but decrease Aß1-42 and p-Tau levels in the hippocampal neurons and increase cell activity. It is suggested that HDAC6 may affect the formation of Aß oligomers and the changes in Tau protein phosphorylation levels in the hippocampus of AD transgenic mouse as well as the alterations in cognitive behavioral functions by regulating the HSP90-HSF1 pathway.

Metformin induction of heat shock factor 1 activation and the mitochondrial unfolded protein response alleviate cardiac remodeling in spontaneously hypertensive rats.

Heart failure and cardiac remodeling are both characterized by mitochondrial dysfunction. Healthy mitochondria are required for adequate contractile activity and appropriate regulation of cell survival. In the mammalian heart, enhancement of the mitochondrial unfolded protein response (UPRmt) is cardioprotective under pressure overload conditions. We explored the UPRmt and the underlying regulatory mechanism in terms of hypertension-induced cardiac remodeling and the cardioprotective effect of metformin. Male spontaneously hypertensive rats and angiotensin II-treated neonatal rat cardiomyocytes were used to induce cardiac hypertrophy. The results showed that hypertension induced the formation of aberrant mitochondria, characterized by a reduced mtDNA/nDNA ratio and swelling, as well as lower levels of mitochondrial complexes I to V and inhibition of the expression of one protein subunit of each of complexes I to IV. Such changes eventually enlarged cardiomyocytes and increased cardiac fibrosis. Metformin treatment increased the mtDNA/nDNA ratio and regulated the UPRmt, as indicated by increased expression of activating transcription factor 5, Lon protease 1, and heat shock protein 60, and decreased expression of C/EBP homologous protein. Thus, metformin improved mitochondrial ultrastructure and function in spontaneously hypertensive rats. In vitro analyses revealed that metformin reduced the high levels of angiotensin II-induced mitochondrial reactive oxygen species in such animals and stimulated nuclear translocation of heat shock factor 1 (HSF1). Moreover, HSF1 small-interfering RNA reduced the metformin-mediated improvements in mitochondrial morphology and the UPRmt by suppressing hypertrophic signals and cardiomyocyte apoptosis. These results suggest that HSF1/UPRmt signaling contributes to the beneficial effects of metformin. Metformin-mediated targeting of mitochondrial protein homeostasis and modulation of HSF1 levels have potential therapeutic implications in terms of cardiac remodeling.

Suppression of NSCLC progression via the co-administration of Danusertib, an AURK inhibitor, and KRIBB11, an HSF1 inhibitor.

Aurora kinase (AURK) and heat shock factor 1 (HSF1) are commonly overexpressed in non-small cell lung cancer (NSCLC), correlating with poor prognosis. This study aims to assess the therapeutic potential of combining the Danusertib (Danu, AURK inhibitor) and KRIBB11 (HSF1 inhibitor) for NSCLC treatment. The effects of this combination were investigated in A549 cells and a tumor xenograft mouse model. The findings demonstrate that concurrent administration of Danu and KRIBB11 effectively impedes cell proliferation, induces apoptosis, and triggers G2/M cell cycle arrest. Moreover, the combination treatment upregulates pro-apoptotic proteins (Cleaved-caspase3, Cleaved-PARP, and Bax) while downregulating anti-apoptotic proteins (Bcl-2), as well as G2/M-related proteins (CDC2 and cyclin B1). Additionally, the combination treatment elevates reactive oxygen species (ROS) levels, decreases mitochondrial membrane potential, and activates the DNA damage pathway. Interestingly, we discovered that the PI3K/AKT pathway is involved in mediating the effects of both Danu and KRIBB11. Furthermore, the combination treatment inhibits tumor growth and AKT signaling in the xenograft mouse model, increases levels of the tumor tissue oxidation product malondialdehyde (MDA), and induces DNA damage. To summarize, a potential therapeutic approach for NSCLC may involve dual inhibition of AURK and HSF1, resulting in the downregulation of the PI3K/AKT signaling pathway, and the activation of ROS-mediated mitochondrial and DNA damage pathways.

MUC1 promotes lymph node metastasis in esophageal squamous cell carcinoma by downregulating DNAJB6 expression.

BACKGROUND: Aberrant expression of MUC1 correlates with the progression of esophageal squamous cell carcinoma (ESCC), this study aimed to explore the effect of targeting MUC1 by Go-203 on malignant behavior of ESCC and the underlying mechanism. METHODS AND RESULTS: IHC was used to examine the expression of MUC1 and DNAJB6 in ESCC samples. qRT-PCR and western blotting were used to examine the expression of MUC1 and DNAJB6 in ESCC cell lines. CCK8, wound healing, and transwell assays were used to determine the effect of regulating MUC1/DNAJB6 on the proliferation, migration, and invasion of ESCC cells. The effect of overexpressing/targeting MUC1 on the activation of the AKT/HSF-1 pathway was determined by western blotting. A negative correlation was confirmed between the expression of DNAJB6 and MUC1 in ESCC tissue samples by IHC, and high expression of MUC1 and low expression of DNAJB6 correlated with lymph node metastasis in ESCC patients. Overexpressing MUC1 downregulated the expression of DNAJB6, promoted ESCC proliferation, invasion, migration and activated the AKT pathway, while targeting MUC1 suppressed proliferation, invasion, migration, and the AKT pathway and up-regulated DNAJB6 expression in vitro. Moreover, MUC1 increased the phosphorylation of HSF-1 via the AKT pathway, and inhibiting AKT-HSF-1 increased the expression of DNAJB6 in vitro. CONCLUSIONS: This study indicated that MUC1 could promote tumorigenesis and metastasis in ESCC by downregulating DNAJB6 expression through AKT-HSF-1 pathway.

Silencing of heat shock factor 1 (HSF1) inhibits proliferation, invasion, and epithelial-mesenchymal transition in oral squamous cell carcinoma.

BACKGROUND: Oral squamous cell carcinoma is characterized by high rates of morbidity and mortality. Evidence obtained for different types of cancer shows that tumor initiation, progression, and therapeutic resistance are regulated by heat shock factor 1. This research aimed to analyze the effects of heat shock factor 1 on the biological behavior of oral squamous cell carcinoma. METHODS: Clinicopathological and immunoexpression study of heat shock factor 1 in 70 cases of oral tongue SCC and functional assays by gene silencing of this factor in an oral tongue SCC cell line. RESULTS: Heat shock factor 1 was overexpressed in oral tongue SCC specimens compared to normal oral mucosa (p < 0.0001) and in the SCC15 line compared to immortalized keratinocytes (p < 0.005). No significant associations were observed between overexpression of heat shock factor 1 and clinicopathological parameters or survival rates of the oral tongue SCC cases in the present sample. In vitro experiments showed that heat shock factor 1 silencing inhibited cell proliferation (p < 0.005) and cell cycle progression, with the accumulation of cells in the G0/G1 phase (p < 0.01). In addition, heat shock factor 1 silencing reduced cell invasion capacity (p < 0.05) and epithelial-mesenchymal transition, characterized by a decrease in vimentin expression (p < 0.05) and an increase in E-cadherin expression (p < 0.001). CONCLUSION: Heat shock factor 1 may exert several functions that help maintain cell stability under the stressful conditions of the tumor microenvironment. Thus, strategies targeting the regulation of this protein may in the future be a useful therapeutic tool to control the progression of oral squamous cell carcinoma.

Monitoring of the Heat Shock Response with a Real-Time Luciferase Reporter.

The heat shock response (HSR) is a cellular mechanism for counteracting acute proteotoxic stress. In eukaryotes, transcriptional activation of the HSR is regulated by heat shock factor 1 (HSF1). Activation of HSF1 induces the expression of heat shock proteins (HSPs) that function as molecular chaperones to fold and maintain the three-dimensional structure of misfolded proteins. The regulation of the degree and duration of the HSR is controlled by multiple biochemical mechanisms that include posttranslational modification of HSF1 and numerous protein-protein interactions. In this chapter, we describe a method to evaluate the activation and deactivation of the HSR at the transcriptional level using a short half-life luciferase reporter assay. This assay can be used to further characterize the HSR or as a screen for small molecule inducers, amplifiers, or repressors.

4-Phenylbutyric acid attenuates amyloid-ß proteotoxicity through activation of HSF-1 in an Alzheimer's disease model of the nematode Caenorhabditiselegans.

Alzheimer's disease (AD) is a neurodegenerative disorder and the most common form of dementia. The pathogenesis is a complex process, in which the proteotoxicity of amyloid-ß (Aß) was identified as a major factor. 4-Phenylbutyric acid (4-PBA) is an aromatic short-chain fatty acid that may attenuate Aß proteotoxicity through its already shown properties as a chemical chaperone or by inhibition of histone deacetylases (HDACs). In the present study, we investigated the molecular effects of 4-PBA on Aß proteotoxicity using the nematode Caenorhabditis elegans as a model. Computer-based analysis of motility was used as a measure of Aß proteotoxicity in the transgenic strain GMC101, expressing human Aß1-42 in body wall muscle cells. Aß aggregation was quantified using the fluorescent probe NIAD-4 to correlate the effects of 4-PBA on motility with the amount of the proteotoxic protein. Furthermore, these approaches were supplemented by gene regulation via RNA interference (RNAi) to identify molecular targets of 4-PBA. 4-PBA improved the motility of GMC101 nematodes and reduced Aß aggregation significantly. Knockdown of hsf-1, encoding an ortholog essential for the cytosolic heat shock response, prevented the increase in motility and decrease in Aß aggregation by 4-PBA incubation. RNAi for hda-1, encoding an ortholog of histone deacetylase 2, also increased motility. Double RNAi for hsf-1 and hda-1 revealed a dominant effect of hsf-1 RNAi. Moreover, 4-PBA failed to further increase motility under hda-1 RNAi. Accordingly, the results suggest that 4-PBA attenuates Aß proteotoxicity in an AD-model of C. elegans through activation of HSF-1 via inhibition of HDA-1.

HSF-1 enhances cardioprotective potential of stem cells via exosome biogenesis and their miRNA cargo enrichment.

Stem cell therapy provides a hope to no option heart disease patient group. Stem cells work via different mechanisms of which paracrine mechanism is reported to justify most of the effects. Therefore, identifying the control arms for paracrine cocktail production is necessary to tailor stem cell functions in disease contextual manner. In this study, we describe a novel paracrine cocktail regulatory axis, in stem cells, to enhance their cardioprotective abilities. We identified that HSF1 knockout resulted in reduced cardiac regenerative abilities of mesenchymal stem cells (MSCs) while its overexpression had opposite effects. Altered exosome biognesis and their miRNA cargo enrichment were found to be underlying these altered regenerative abilities. Decreased production of exosomes by MSCs accompanied their loss of HSF1 and vice versa. Moreover, the exosomes derived from HSF1 depleted MSCs showed significantly reduced candidate miRNA expression (miR-145, miR-146, 199-3p, 199b and miR-590) compared to those obtained from HSF1 overexpressing MSCs. We further discovered that HSF1 mediates miRNAs' enrichment into exosomes via Y binding protein 1 (YBX1) and showed, by loss and gain of function strategies, that miRNAs' enrichment in mesenchymal stem cell derived exosomes is deregulated with altered YBX1 expression. It was finally demonstrated that absence of YBX1 in MSCs, with normal HSF1 expression, resulted in significant accumulation of candidate miRNAs into the cells. Together, our data shows that HSF1 plays a critical role in determining the regenerative potential of stem cells. HSF1 does that by affecting exosome biogenesis and miRNA cargo sorting via regulation of YBX1 gene expression.

Heat shock factor 1 is a promising therapeutic target against adult T-cell leukemia.

Patients with adult T-cell leukemia (ATL), which is caused by human T-cell leukemia virus type 1 (HTLV-1), show poor prognosis because of drug resistance. Heat shock protein (HSP) 90 is reportedly essential for ATL cell survival as it regulates important signaling pathways, thereby making HSP90 inhibitors new therapeutic candidates for ATL. However, HSP90 inhibition increases the expression of other HSPs, suggesting that HSPs may contribute to drug resistance. The heat shock factor 1 (HSF1) transcription factor is the primary regulator of the expression of HSPs. Furthermore, targeting HSF1 disrupts the HSP90 chaperone function. Herein, we demonstrated that HSF1 is overexpressed in HTLV-1-infected T cells. HSF1 knockdown inhibited the proliferation of HTLV-1-infected T cells. HSF1 inhibitor KRIBB11 reduced the expression and phosphorylation of HSF1, downregulated HSP70 and HSP27 expression, and suppressed Akt, nuclear factor-κB, and AP-1 signals. KRIBB11 treatment induced DNA damage, upregulated p53 and p21, and reduced the expression of cyclin D2/E, CDK2/4, c-Myc, MDM2, and ß-catenin, thereby preventing retinoblastoma protein phosphorylation and inhibiting G1-S cell cycle progression. KRIBB11 also induced caspase-mediated apoptosis concomitant with the suppression of Bcl-xL, Mcl-1, XIAP, c-IAP1/2, and survivin expression. KRIBB11 inhibited HSP70 and HSP90 upregulation through treatment with AUY922, an HSP90 inhibitor, and enhanced the cytotoxic effect of AUY922, suggesting a salvage role of HSF1-dependent HSP induction in response to drug treatment. Finally, treatment of mice with KRIBB11 reduced ATL tumor growth. Therefore, this study provides a strong rationale to target HSF1 and validates the anti-ATL activity of KRIBB11.

Proteotoxic stress response in atherosclerotic cardiovascular disease: Emerging role of heat shock factor 1.

Atherosclerosis is a major risk factor for cardiovascular diseases. Hypercholesterolemia has been both clinically and experimentally linked to cardiovascular disease and is involved in the initiation of atherosclerosis. Heat shock factor 1 (HSF1) is involved in the control of atherosclerosis. HSF1 is a critical transcriptional factor of the proteotoxic stress response that regulates the production of heat shock proteins (HSPs) and other important activities such as lipid metabolism. Recently, HSF1 is reported to directly interact with and inhibit AMP-activated protein kinase (AMPK) to promote lipogenesis and cholesterol synthesis. This review highlights roles of HSF1 and HSPs in critical metabolic pathways of atherosclerosis, including lipogenesis and proteome homeostasis.

Heat shock factor 1 promotes neurite outgrowth and suppresses inflammation in the severed spinal cord of geckos.

The low intrinsic growth capacity of neurons and an injury-induced inhibitory milieu are major contributors to the failure of sensory and motor functional recovery following spinal cord injury. Heat shock transcription factor 1 (HSF1), a master regulator of the heat shock response, plays neurogenetic and neuroprotective roles in the damaged or diseased central nervous system. However, the underlying mechanism has not been fully elucidated. In the present study, we used a gecko model of spontaneous nerve regeneration to investigate the potential roles of gecko HSF1 (gHSF1) in the regulation of neurite outgrowth and inflammatory inhibition of macrophages following spinal cord injury. gHSF1 expression in neurons and microglia at the lesion site increased dramatically immediately after tail amputation. gHSF1 overexpression in gecko primary neurons significantly promoted axonal growth by suppressing the expression of suppressor of cytokine signaling-3, and facilitated neuronal survival via activation of the mitogen-activated extracellular signal-regulated kinase/extracellular regulated protein kinases and phosphatidylinositol 3-kinase/protein kinase B pathways. Furthermore, gHSF1 efficiently inhibited the macrophage-mediated inflammatory response by inactivating IkappaB-alpha/NF-kappaB signaling. Our findings show that HSF1 plays dual roles in promoting axonal regrowth and inhibiting leukocyte inflammation, and provide new avenues of investigation for promoting spinal cord injury repair in mammals.

Activation of HSP90/HSF1 Signaling as an Adaptive Response to an Electrophilic Metabolite of Morphine.

Morphinone (MO) is an electrophilic metabolite of morphine that covalently binds to protein thiols, resulting in toxicity in vitro and in vivo. We have previously identified a variety of redox signaling pathways that are activated during electrophilic stress. However, the role of MO in such activation remains unknown. In this study, we examined whether MO could activate heat shock protein (HSP) 90/heat shock factor (HSF) 1 signaling in HepG2 cells. MO exposure caused S-modification of HSP90 (determined using biotin-PEAC5-maleimide labeling) and nuclear translocation of transcription factor HSF1, thereby up-regulating its downstream genes encoding B-cell lymphoma 2-associated anthanogene 3 and heat shock 70 kDa protein 1. However, dihydromorphinone, a non-electrophilic metabolite of morphine, had little effect on HSF1 activation or upregulation of these genes, suggesting that covalent modification plays a role in this process and that the HSP90/HSF1 pathway is a redox-signaled adaptive response to morphine metabolism.

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.

HSF1 Attenuates the Release of Inflammatory Cytokines Induced by Lipopolysaccharide through Transcriptional Regulation of Atg10.

Autophagy plays an important role in endotoxemic mice, and heat shock factor 1 (HSF1) plays a crucial protective role in endotoxemic mice. However, the protective mechanisms of HSF1 are poorly understood. In this text, bioinformatics analysis, chromatin immunoprecipitation, and electrophoresis mobility shift assay were employed to investigate the underlying mechanisms. The results showed that the release of inflammatory cytokines increased and autophagy decreased significantly in Hsf1-/- endotoxemic mice compared with those in Hsf1+/+ endotoxemic mice. HSF1 could directly bind to the noncoding promoter region of the autophagy-related gene 10 (Atg10). The expression of ATG10 and the ratio of LC3-II/LC3-I were obviously decreased in LPS-treated Hsf1-/- peritoneal macrophages (PM) versus those in LPS-treated Hsf1+/+ PM. Overexpression of HSF1 increased the level of the ATG10 protein and enhanced the ratio of LC3-II/LC3-I in RAW264.7 cells. In contrast, silencing of HSF1 decreased the expression of ATG10 and markedly lowered the ratio of LC3-II/LC3-I. In a cotransfected cell experiment, the upregulation of autophagy by overexpression HSF1 was reversed by small interfering RNA (siRNA)-ATG10. Compared with the overexpression HSF1, the release of inflammatory cytokines induced by lipopolysaccharide (LPS) was decreased in pcDNA3.1-HSF1 with siRNA-ATG10 cotransfected RAW264.7 cells. On the other hand, the decrease of autophagy by siRNA-HSF1 was compensated by overexpression of ATG10. Compared with siRNA-HSF1, the release of inflammatory cytokines induced by LPS was increased in siRNA-HSF1 with pcDNA3.1-ATG10 cotransfected RAW264.7 cells. These results presented a novel mechanism that HSF1 attenuated the release of inflammatory cytokines induced by LPS through transcriptional regulation of Atg10. Targeting of HSF1-Atg10-autophagy might be an attractive strategy in endotoxemia therapeutics. IMPORTANCE HSF1 plays an important protective role in endotoxemic mice. However, the protective mechanisms of HSF1 are poorly understood. In the present study, we demonstrated that HSF1 upregulated ATG10 through specifically binding Atg10 promoter's noncoding region in LPS-treated PM and RAW264.7 cells. By depletion of HSF1, the expression of ATG10 was significantly decreased, leading to aggravate releasing of inflammatory cytokines in LPS-treated RAW264.7 cells. These findings provided a new mechanism of HSF1 in endotoxemic mice.

HSF1, Aging, and Neurodegeneration.

Heat shock factor 1 (HSF1) is a master transcription regulator that mediates the induction of heat shock protein chaperones for quality control (QC) of the proteome and maintenance of proteostasis as a protective mechanism in response to stress. Research in this particular area has accelerated dramatically over the past three decades following successful isolation, cloning, and characterization of HSF1. The intricate multi-protein complexes and transcriptional activation orchestrated by HSF1 are fundamental processes within the cellular QC machinery. Our primary focus is on the regulation and function of HSF1 in aging and neurodegenerative diseases (ND) which represent physiological and pathological states of dysfunction in protein QC. This chapter presents an overview of HSF1 structural, functional, and energetic properties in healthy cells while addressing the deterioration of HSF1 function viz-à-viz age-dependent and neuron-specific vulnerability to ND. We discuss the structural domains of HSF1 with emphasis on the intrinsically disordered regions and note that disease proteins associated with ND are often structurally disordered and exquisitely sensitive to changes in cellular environment as may occur during aging. We propose a hypothesis that age-dependent changes of the intrinsically disordered proteome likely hold answers to understand many of the functional, structural, and organizational changes of proteins and signaling pathways in aging - dysfunction of HSF1 and accumulation of disease protein aggregates in ND included.Structured AbstractsIntroduction: Heat shock factor 1 (HSF1) is a master transcription regulator that mediates the induction of heat shock protein chaperones for quality control (QC) of the proteome as a cyto-protective mechanism in response to stress. There is cumulative evidence of age-related deterioration of this QC mechanism that contributes to disease vulnerability. OBJECTIVES: Herein we discuss the regulation and function of HSF1 as they relate to the pathophysiological changes of protein quality control in aging and neurodegenerative diseases (ND). METHODS: We present an overview of HSF1 structural, functional, and energetic properties in healthy cells while addressing the deterioration of HSF1 function vis-à-vis age-dependent and neuron-specific vulnerability to neurodegenerative diseases. RESULTS: We examine the impact of intrinsically disordered regions on the function of HSF1 and note that proteins associated with neurodegeneration are natively unstructured and exquisitely sensitive to changes in cellular environment as may occur during aging. CONCLUSIONS: We put forth a hypothesis that age-dependent changes of the intrinsically disordered proteome hold answers to understanding many of the functional, structural, and organizational changes of proteins - dysfunction of HSF1 in aging and appearance of disease protein aggregates in neurodegenerative diseases included.