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Cisplatin is a commonly used chemotherapy agent with a nearly universal side effect of sensorineural hearing loss. The cellular mechanisms underlying cisplatin ototoxicity are poorly understood. Efforts in drug development to prevent or reverse cisplatin ototoxicity have largely focused on pathways of oxidative stress and apoptosis. An effective treatment for cisplatin ototoxicity, sodium thiosulfate (STS), while beneficial when used in standard risk hepatoblastoma, is associated with reduced survival in disseminated pediatric malignancy, highlighting the need for more specific drugs without potential tumor protective effects. The unfolded protein response (UPR) and endoplasmic reticulum (ER) stress pathways have been shown to be involved in the pathogenesis of noise-induced hearing loss and cochlear synaptopathy in vivo, and these pathways have been implicated broadly in cisplatin cytotoxicity. This study sought to determine whether the UPR can be targeted to prevent cisplatin ototoxicity. Neonatal cochlear cultures and HEK cells were exposed to cisplatin, and UPR marker gene expression and cell death measured. Treatment with ISRIB (Integrated Stress Response InhIBitor), a drug that activates eif2B and downregulates the pro-apoptotic PERK/CHOP pathway of the UPR, was tested for its ability to reduce apoptosis in HEK cells, hair-cell death in cochlear cultures, and hearing loss using an in vivo mouse model of cisplatin ototoxicity. Finally, to evaluate whether ISRIB might interfere with cisplatin chemoeffectiveness, we tested it in head and neck squamous cell carcinoma (HNSCC) cell-based assays of cisplatin cytotoxicity. Cisplatin exhibited a biphasic, non-linear dose-response of cell death and apoptosis that correlated with different patterns of UPR marker gene expression in HEK cells and cochlear cultures. ISRIB treatment protected against cisplatin-induced hearing loss and hair-cell death, but did not impact cisplatins cytotoxic effects on HNSCC cell viability, unlike STS. These findings demonstrate that targeting the pro-apoptotic PERK/CHOP pathway with ISRIB can mitigate cisplatin ototoxicity without reducing anti-cancer cell effects, suggesting that this may be a viable strategy for drug development.

The central nervous system coordinates peripheral cellular stress responses, including the unfolded protein response of the mitochondria (UPRMT); however, the contexts for which this regulatory capability evolved are unknown. UPRMT is up-regulated upon pathogenic infection and in metabolic flux, and the olfactory nervous system has been shown to regulate pathogen resistance and peripheral metabolic activity. Therefore, we asked whether the olfactory nervous system in Caenorhabditis elegans controls the UPRMT cell nonautonomously. We found that silencing a single inhibitory olfactory neuron pair, AWC, led to robust induction of UPRMT and reduction of oxidative phosphorylation dependent on serotonin signaling and parkin-mediated mitophagy. Further, AWC ablation confers resistance to the pathogenic bacteria Pseudomonas aeruginosa partially dependent on the UPRMT transcription factor atfs-1 and fully dependent on mitophagy machinery. These data illustrate a role for the olfactory nervous system in regulating whole-organism mitochondrial dynamics, perhaps in preparation for postprandial metabolic stress or pathogenic infection.

The endoplasmic reticulum (ER) is a single-copy organelle that cannot be generated de novo, suggesting coordination between the mechanisms overseeing ER integrity and those controlling the cell cycle to maintain organelle inheritance. The Unfolded Protein Response (UPR) is a conserved signaling network that regulates ER homeostasis. Here, we show that pharmacological and genetic inhibition of the UPR sensors IRE1, ATF6, and PERK in unstressed cells delays the cell cycle, with PERK inhibition showing the most penetrant effect, which was associated with a slowdown of the G1-to-S/G2 transition. Treatment with the small molecule ISRIB to bypass the effects of PERK-dependent phosphorylation of the translation initiation factor eIF2α had no such effect, suggesting that cell cycle timing depends on PERKs kinase activity but is independent of eIF2α phosphorylation. Using complementary light and electron microscopy and flow cytometry-based analyses, we also demonstrate that the ER enlarges before mitosis. Together, our results suggest coordination between UPR signaling and the cell cycle to maintain ER physiology during cell division.

Aquaporin-1 (Aqp1), a water channel, has garnered significant interest for cell-based medicine and in vivo synthetic biology due to its ability to be genetically encoded to produce magnetic resonance signals by increasing the rate of water diffusion in cells. However, concerns regarding the effects of Aqp1 overexpression and increased membrane diffusivity on cell physiology have limited its widespread use as a deep-tissue reporter. In this study, we present evidence that Aqp1 generates strong diffusion-based magnetic resonance signals without adversely affecting cell viability or morphology in diverse cell lines derived from mice and humans. Our findings indicate that Aqp1 overexpression does not induce ER stress, which is frequently associated with heterologous expression of membrane proteins. Furthermore, we observed that Aqp1 expression had no detrimental effects on native biological activities, such as phagocytosis, immune response, insulin secretion, and tumor cell migration in the analyzed cell lines. These findings should serve to alleviate any lingering safety concerns regarding the utilization of Aqp1 as a genetic reporter and should foster its broader application as a noninvasive reporter for in vivo studies.

Friedreich ataxia (FRDA) is a multisystemic, autosomal recessive disorder caused by homozygous GAA expansion mutation in the first intron of frataxin (FXN) gene. FXN is a mitochondrial protein critical for iron-sulfur cluster biosynthesis and deficiency impairs mitochondrial electron transport chain functions and iron homeostasis within the organelle. Currently, there is no effective treatment for FRDA. We have previously demonstrated that single infusion of wild-type hematopoietic stem and progenitor cells (HSPCs) resulted in prevention of neurologic and cardiac complications of FRDA in YG8R mice, and rescue was mediated by FXN transfer from tissue engrafted, HSPC-derived microglia/macrophages to diseased neurons/myocytes. For a future clinical translation, we developed an autologous stem cell transplantation approach using CRISPR/Cas9 for the excision of the GAA repeats in FRDA patients CD34+ HSPCs; this strategy leading to increased FXN expression and improved mitochondrial functions. The aim of the current study is to validate the efficiency and safety of our gene editing approach in a disease-relevant model. We generated a cohort of FRDA patient-derived iPSCs and isogenic lines that were gene edited with our CRISPR/Cas9 approach. iPSC derived FRDA neurons displayed characteristic apoptotic and mitochondrial phenotype of the disease, such as non-homogenous microtubule staining in neurites, increased caspase-3 expression, mitochondrial superoxide levels, mitochondrial fragmentation, and partial degradation of the cristae compared to healthy controls. These defects were fully prevented in the gene edited neurons. RNASeq analysis of FRDA and gene edited neurons demonstrated striking improvement in gene clusters associated with endoplasmic reticulum (ER) stress in the isogenic lines. Gene edited neurons demonstrated improved ER-calcium release, normalization of ER stress response gene, XBP-1, and significantly increased ER-mitochondrial contacts that are critical for functional homeostasis of both organelles, as compared to FRDA neurons. Ultrastructural analysis for these contact sites displayed severe ER structural damage in FRDA neurons, that was undetected in gene edited neurons. Taken together, these results represent a novel finding for disease pathogenesis showing dramatic ER structural damage in FRDA, validate the efficacy profile of our FXN gene editing approach in a disease relevant model, and support our approach as an effective strategy for therapeutic intervention for Friedreichs ataxia.

Background: Colorectal cancer (CRC) is the most common gastrointestinal malignancy worldwide, with obesity-induced lipid metabolism disorders playing a crucial role in its progression. A complex connection exists between gut microbiota and the development of intestinal tumors through the microbiota metabolite pathway. Metabolic disorders frequently alter the gut microbiome, impairing immune and cellular functions and hastening cancer progression. Methods: This study thoroughly examined the gut microbiota through 16S rRNA sequencing of fecal samples from 181 CRC patients, integrating preoperative Low-density lipoprotein cholesterol (LDL-C) levels and RNA sequencing data. The study includes a comparison of microbial diversity, differential microbiological analysis, exploration of the associations between microbiota, tumor microenvironment immune cells, and immune genes, enrichment analysis of potential biological functions of microbe-related host genes, and the prediction of LDL-C status through microorganisms. Results: The analysis revealed that differences in α and β diversity indices of intestinal microbiota in CRC patients were not statistically significant across different LDL-C metabolic states. Patients exhibited varying LDL-C metabolic conditions, leading to a bifurcation of their gut microbiota into two distinct clusters. Patients with LDL-C metabolic irregularities had higher concentrations of twelve gut microbiota, which were linked to various immune cells and immune-related genes, influencing tumor immunity. Under normal LDL-C metabolic conditions, the protective microorganism Anaerostipes_caccae was significantly negatively correlated with the GO Biological Process pathway involved in the negative regulation of the unfolded protein response in the endoplasmic reticulum. Both XGBoost and MLP models, developed using differential gut microbiota, could forecast LDL-C levels in CRC patients biologically. Conclusions: The intestinal microbiota in CRC patients influences the LDL-C metabolic status. With elevated LDL-C levels, gut microbiota can regulate the function of immune cells and gene expression within the tumor microenvironment, affecting cancer-related pathways and promoting CRC progression. LDL-C and its associated gut microbiota could provide non-invasive markers for clinical evaluation and treatment of CRC patients. [ABSTRACT FROM AUTHOR]

Introduction: The contribution of Cannabinoid type 1 receptor (CB1) in mitochondrial energy transduction mechanisms and mitochondrial activities awaits deeper investigations. Our study aims to assess the impact of CB1 absence on the mitochondrial compartment in the liver, focusing on both functional aspects and remodeling processes. Methods: We used CB1−/− and CB1+/+ male mice. Cytochrome C Oxidase activity was determined polarographically. The expression and the activities of separated mitochondrial complexes and supercomplexes were performed by using Blue-Native Page, Western blotting and histochemical staining for in-gel activity. Key players of Mitochondrial Quality Control processes were measured using RT-qPCR and Western blotting. Liver fine sub-cellular ultrastructural features were analyzed by TEM analysis. Results and discussion: In the absence of CB1, several changes in the liver occur, including increased oxidative capacity, reduced complex I activity, enhanced complex IV activity, general upregulation of respiratory supercomplexes, as well as higher levels of oxidative stress. The mitochondria and cellular metabolism may be affected by these changes, increasing the risk of ROS-related damage. CB1−/− mice show upregulation of mitochondrial fusion, fission and biogenesis processes which suggests a dynamic response to the absence of CB1. Furthermore, oxidative stress disturbs mitochondrial proteostasis, initiating the mitochondrial unfolded protein response (UPRmt). We noted heightened levels of pivotal enzymes responsible for maintaining mitochondrial integrity, along with heightened expression of molecular chaperones and transcription factors associated with cellular stress reactions. Additionally, our discoveries demonstrate a synchronized reaction to cellular stress, involving both UPRmt and UPRER pathways. [ABSTRACT FROM AUTHOR]

ER-phagy, a selective form of autophagic degradation of endoplasmic reticulum (ER) fragments, plays an essential role in governing ER homeostasis. Dysregulation of ER-phagy is associated with the unfolded protein response (UPR), which is a major clue for evoking inflammatory diseases. However, the molecular mechanism underpinning the connection between ER-phagy and disease remains poorly defined. Here, we identified ubiquitin-associated domain-containing protein 2 (UBAC2) as a receptor for ER-phagy, while at the same time being a negative regulator of inflammatory responses. UBAC2 harbors a canonical LC3-interacting region (LIR) in its cytoplasmic domain, which binds to autophagosomal GABARAP. Upon ER-stress or autophagy activation, microtubule affinity-regulating kinase 2 (MARK2) phosphorylates UBAC2 at serine (S) 223, promoting its dimerization. Dimerized UBAC2 interacts more strongly with GABARAP, thus facilitating selective degradation of the ER. Moreover, by affecting ER-phagy, UBAC2 restrains inflammatory responses and acute ulcerative colitis (UC) in mice. Our findings indicate that ER-phagy directed by a MARK2-UBAC2 axis may provide targets for the treatment of inflammatory disease. Synopsis: Endoplasmic reticulum (ER) homeostasis is critical for maintaining optimal host immune responses. This study identifies UBAC2 as a phospho-regulated ER-phagy receptor that can suppress the unfolded protein response and sterile inflammation. UBAC2 is an ER resident protein that undergoes autophagic degradation upon binding GABARAP. UBAC2 promotes ER-phagy flux through its LC3-interacting region (LIR). MARK2 phosphorylates UBAC2 at serine 223, resulting in the dimerization of UBAC2 to enhance the ER-phagy process. A somatic mutation near the LIR motif of UBAC2 increases ER stress responses and sterile inflammation. Phosphorylation-induced UBAC2 dimerization facilitates binding to autophagosomal GABARAP and restrains acute ulcerative colitis via ER-phagy. [ABSTRACT FROM AUTHOR]

Bronchopulmonary dysplasia (BPD) is the most common complication of prematurity. Oxidative stress (OS) and inflammation are the major contributors to BPD. Despite aggressive treatments, BPD prevalence remains unchanged, which underscores the urgent need to explore more potential therapies. The endoplasmic reticulum (ER) plays crucial roles in surfactant and protein synthesis, assisting mitochondrial function, and maintaining metabolic homeostasis. Under OS, disturbed metabolism and protein folding transform the ER structure to refold proteins and help degrade non-essential proteins to resume cell homeostasis. When OS becomes excessive, the endogenous chaperone will leave the three ER stress sensors to allow subsequent changes, including cell death and senescence, impairing the growth potential of organs. The contributing role of ER stress in BPD is confirmed by reproducing the BPD phenotype in rat pups by ER stress inducers. Although chemical chaperones attenuate BPD, ER stress is still associated with cellular senescence. N-acetyl-lysyltyrosylcysteine amide (KYC) is a myeloperoxidase inhibitor that attenuates ER stress and senescence as a systems pharmacology agent. In this review, we describe the role of ER stress in BPD and discuss the therapeutic potentials of chemical chaperones and KYC, highlighting their promising role in future therapeutic interventions. [ABSTRACT FROM AUTHOR]

Any opacification of the lens can be defined as cataracts, and lens epithelium cells play a crucial role in guaranteeing lens transparency by maintaining its homeostasis. Although several causative genes of congenital cataracts have been reported, the mechanisms underlying lens opacity remain unclear. In this study, a large family with congenital cataracts was collected and genetic analysis revealed a pathological mutation (c.3857 C > T, p.T1287I) in the GBF1 gene; all affected individuals in the family carried this heterozygous mutation, while unaffected family members did not. Functional studies in human lens epithelium cell line revealed that this mutation led to a reduction in GBF1 protein levels. Knockdown of endogenous GBF1 activated XBP1s in the unfolded protein response signal pathway, and enhances autophagy in an mTOR-independent manner. Heterozygous Gbf1 knockout mice also displayed typic cataract phenotype. Together, our study identified GBF1 as a novel causative gene for congenital cataracts. Additionally, we found that GBF1 deficiency activates the unfolded protein response and leads to enhanced autophagy, which may contribute to lens opacity. [ABSTRACT FROM AUTHOR]

Introduction: Colorectal cancer ranks as the third most common cancer in both men and women, with approximately 35% of cases being stage IV metastatic at diagnosis. Even with treatment advancements, the survival rates for these patients remain suboptimal. There is a significant focus on developing multi-targeted therapies due to the common issue of drug resistance in standard and targeted cancer treatments. Medicinal mushrooms, both as single compounds and as complex extracts, have undergone extensive research. Numerous types of mushrooms have been shown to be safe, effective inhibitors of cancer pathways and strong enhancers of the immune system. Methods: In this study, we performed both qualitative and quantitative proteomic analyses using tandem mass tags (TMT) on CT26 wild type (CT26. WT) colon cancer tissues from Balb/c mice, which were treated with a special blend of medicinal mushroom extracts, either alone or in combination with the chemotherapy drug 5-fluorouracil. Results: The results showed a notable increase in survival rates and indicated that medicinal mushroom preparation Agarikon Plus, both alone and combined with 5-fluorouracil or another medicinal mushroom preparation Agarikon.1, impedes multiple key processes in colorectal cancer progression. The analysis of differentially expressed proteins in treated groups was done by use of bioinformatics tools and a decrease in ribosomal biogenesis (e.g., RPS3) and translation processes (e.g., RPL14) as well as an increase in unfolded protein response (e.g., DNAJC3), lipid metabolism (e.g., ACOT7), and the tricarboxylic acid cycle (e.g., FH) were observed. Conclusion: The treatment induced various alterations of known biomarkers and protein clusters critical to the progression and prognosis of colorectal cancer, laying a promising foundation for further translational research on this treatment modality. [ABSTRACT FROM AUTHOR]

Bladder cancer stands as one of the most prevalent cancers worldwide. While our previous research confirmed the significant role of stearoyl-CoA desaturase (SCD) in bladder cancer, the underlying reasons for its abnormal overexpression remain largely unknown. Moreover, the distinct response to SCD inhibitors between cancer stem cells (CSCs) and adherent cultured cell lines lacks clear elucidation. Therefore, in this experiment, we aim to conduct an analysis and screening of the SCD transcription start site, further seeking critical transcription factors involved. Simultaneously, through experimental validation, we aim to explore the pivotal role of endoplasmic reticulum stress/unfolded protein response in drug sensitivity among cancer stem cells. Additionally, our RNA-seq and lipid metabolism analysis revealed the significant impact of nervonic acid on altering the proliferative capacity of bladder cancer cell lines. [ABSTRACT FROM AUTHOR]

Objective: Chemotherapy during pregnancy has a certain risk of causing a series of complications, such as miscarriage, premature birth, or fetal growth restriction, although the relationship between these complications and chemotherapy is currently unclear. This experiment focuses on the possible damage mechanism of the chemotherapeutic drug paclitaxel on placental trophoblast cells, and explores whether chemotherapy can affect pregnancy outcomes by directly damaging placental tissue. Methods: This study explored the mechanism of paclitaxel induced damage on placental trophoblast cell lines JEG-3 and BEWO through immunofluorescence staining, Western blot experiments, cell flow cytometry, Seahorese cell metabolism experiments, and mouse modeling verification. Results: The experiment found that paclitaxel could induce JEG-3 and BEWO cells to produce reactive oxygen species (ROS), and elevate the ratio of Bax/Bcl-2 expression. Besides, paclitaxel mediated the reduction of mitochondrial membrane potential in JEG-3 and BEWO cells, causing damage and leading to mitochondrial autophagy and the occurrence of unfolded protein response. Paclitaxel inhibited the glycolysis rate of JEG-3 and BEWO cells, and leaded to impaired mitochondrial function, including decreased basal respiratory values, decreased respiratory reserve capacity, and proton leakage. In pregnant mice with tumor modeling, paclitaxel could cause DNA damage in placental tissue cells, and might lead to apoptosis of chemotherapy mice placental tissue cells and impairment of normal physiological functions. Conclusion: Paclitaxel may directly or indirectly affect the normal physiological functions of placental trophoblast cells, including energy metabolism and protein synthesis dysfunction, which may be related to the adverse pregnancy outcomes caused by paclitaxel chemotherapy. [ABSTRACT FROM AUTHOR]

The Endoplasmic Reticulum (ER)-resident HSP70 chaperone BiP (HSPA5) plays a crucial role in maintaining and restoring protein folding homeostasis in the ER. BiP's function is often dysregulated in cancer and virus-infected cells, conferring pro-oncogenic and pro-viral advantages. We explored BiP's functions during infection by the Kaposi's sarcoma-associated herpesvirus (KSHV), an oncogenic gamma-herpesvirus associated with cancers of immunocompromised patients. Our findings reveal that BiP protein levels are upregulated in infected epithelial cells during the lytic phase of KSHV infection. This upregulation occurs independently of the unfolded protein response (UPR), a major signaling pathway that regulates BiP availability. Genetic and pharmacological inhibition of BiP halts KSHV viral replication and reduces the proliferation and survival of KSHV-infected cells. Notably, inhibition of BiP limits the spread of other alpha- and beta-herpesviruses and poxviruses with minimal toxicity for normal cells. Our work suggests that BiP is a potential target for developing broad-spectrum antiviral therapies against double-stranded DNA viruses and a promising candidate for therapeutic intervention in KSHV-related malignancies. Author summary: The endoplasmic reticulum (ER) chaperone protein BiP (HSPA5) plays a central role in protein folding and maintaining homeostasis within the ER. Under certain conditions, such as cancer and viral infections, BiP is dysregulated to support cell survival or viral replication. In this study, we investigated the regulation and requirement of BiP during infection by Kaposi's sarcoma-associated herpesvirus (KSHV), an oncogenic herpesvirus linked to cancers in immunocompromised individuals. Our findings demonstrate that BiP is significantly upregulated in KSHV-infected cells, even when the expression of most other cellular genes is suppressed. Notably, the function of BiP is essential for KSHV replication and the survival of KSHV-infected cells. This reliance on BiP is not unique to KSHV; it is also required for the replication of other double-stranded DNA (dsDNA) viruses, including Herpes simplex virus I, Human Cytomegalovirus, and Vaccinia Virus. These findings underscore the critical role of BiP in dsDNA viral infections, positioning this chaperone as a promising target for the development of broad-spectrum antiviral therapies and potential treatment strategies for KSHV-associated malignancies. [ABSTRACT FROM AUTHOR]

Background: The transcriptomic profile of cellular senescence is strongly associated with distinct cell types, the specific stressors triggering senescence, and temporal progression through senescence stages. This implies the potential necessity of conducting separate investigations for each cell type and a stressor inducing senescence. To elucidate the molecular mechanism that drives endoplasmic reticulum (ER) stress-induced cellular senescence in MCF-7 breast cancer cells, with a particular emphasis on the ATF6α branch of the unfolded protein response. We conducted transcriptomic analysis on MCF-7 cells by ectopic expression of ATF6α. Methods: Transcriptomic sequencing was conducted on MCF-7 cells at 6 and 9 hours post senescence induction through ATF6α ectopic expression. Comprehensive analyses encompassing enriched functional annotation, canonical pathway analysis, gene network analysis, upstream regulator analysis and gene set enrichment analysis were performed on Differentially Expressed Genes (DEGs) at 6 and 9 hours as well as time-related DEGs. Regulators and their targets identified from the upstream regulator analysis were validated through RNA interference, and their impact on cellular senescence was assessed by senescence-associated β-galactosidase staining. Results: ATF6α ectopic expression resulted in the identification of 12 and 79 DEGs at 6 and 9 hours, respectively, employing criteria of a false discovery rate < 0.05 and a lower fold change (FC) cutoff |log2FC| > 1. Various analyses highlighted the involvement of the UPR and/or ER Stress Pathway. Upstream regulator analysis of 9 hour-DEGs identified six regulators and eleven target genes associated with processes related to cytostasis and 'cell viability and cell death of connective tissue cells.' Validation confirmed the significance of MAP2K1/2, GPAT4, and PDGF-BB among the regulators and DDIT3, PPP1R15A, and IL6 among the targets. Conclusion: Transcriptomic analyses and validation reveal the importance of the MAP2K1/2/GPAT4-DDIT3 pathway in driving cellular senescence following ATF6α ectopic expression in MCF-7 cells. This study contributes to our understanding of the initial molecular events underlying ER stress-induced cellular senescence in breast cancer cells, providing a foundation for exploring cell type- and stressor-specific responses in cellular senescence induction. [ABSTRACT FROM AUTHOR]

Glioblastoma is a highly aggressive brain tumor for which there is no cure. The dire prognosis of this disease is largely attributable to a high level of heterogeneity, including the presence of a subpopulation of tumor‐initiating glioblastoma stem‐like cells (GSCs), which are refractory to chemo‐ and radiotherapy. Here, in an unbiased marine‐derived fungal extract screen, together with bioguided dereplication based on high‐resolution mass spectrometry, we identified malformin C to preferentially induce cell death in patient‐derived GSCs and explore the potential of this cyclic peptide as a therapeutic agent for glioblastoma. Malformin C significantly reduced tumor growth in an in vivo xenograft model of glioblastoma. Using transcriptomics and chemoproteomics, we found that malformin C binds to many proteins, leading to their aggregation, and rapidly induces the unfolded protein response, including autophagy, in GSCs. Crucially, chemical inhibition of translation using cycloheximide rescued malformin C‐induced cell death in GSCs, demonstrating that the proteotoxic effect of the compound is necessary for its cytotoxicity. At the same time, malformin C appears to accumulate in lysosomes, disrupting autophagic flux, and driving cells to death. Supporting this, malformin C synergizes with chloroquine, an inhibitor of autophagy. Strikingly, we observed that autophagic flux is differentially regulated in GSCs compared with normal astrocytes. The sensitivity of GSCs to malformin C highlights the relevance of proteostasis and autophagy as a therapeutic vulnerability in glioblastoma. [ABSTRACT FROM AUTHOR]

Regulating cardiolipin to maintain mitochondrial homeostasis is a promising strategy for addressing Parkinson's disease (PD). Through a comprehensive screening and validation process involving multiple models, ginsenoside Rg3 (Rg3) as a compound capable of enhancing cardiolipin levels is identified. This augmentation in cardiolipin levels fosters mitochondrial homeostasis by bolstering mitochondrial unfolded protein response, promoting mitophagy, and enhancing mitochondrial oxidative phosphorylation. Consequently, this cascade enhances the survival of tyrosine hydroxylase positive (TH+) dopaminergic neurons, leading to an amelioration in motor performance within PD mouse models. Using limited proteolysis–small‐molecule mapping combined with molecular docking analysis, it has confirmed Growth Factor Receptor‐Bound Protein 2 (GRB2) as a molecular target for Rg3. Furthermore, these investigations reveal that Rg3 facilitates the interaction between GRB2 and TRKA (Neurotrophic Tyrosine Kinase, Receptor, Type 1), thus promotes EVI1 (Ecotropic Virus Integration Site 1 Protein Homolog) phosphorylation by ERK, subsequently increases CRLS1 (Cardiolipin Synthase 1) gene expression and boosts cardiolipin synthesis. The absence of GRB2 or CRLS1 significantly attenuates the beneficial effects of Rg3 on PD symptoms. Finally, Tenofovir Disoproxil Fumarate (TDF) that also promotes the binding between GRB2 and TRKA is further identified. The identified compounds, Rg3 and TDF, exhibit promising potential for the prevention of PD by bolstering cardiolipin expression and reinstating mitochondrial homeostasis. [ABSTRACT FROM AUTHOR]

Canonical models of intestinal regeneration emphasize the critical role of the crypt stem cell niche to generate enterocytes that migrate to villus ends. Burmese pythons possess extreme intestinal regenerative capacity yet lack crypts, thus providing opportunities to identify noncanonical but potentially conserved mechanisms that expand our understanding of regenerative capacity in vertebrates, including humans. Here, we leverage single-nucleus RNA sequencing of fasted and postprandial python small intestine to identify the signaling pathways and cell–cell interactions underlying the python’s regenerative response. We find that python intestinal regeneration entails the activation of multiple conserved mechanisms of growth and stress response, including core lipid metabolism pathways and the unfolded protein response in intestinal enterocytes. Our single-cell resolution highlights extensive heterogeneity in mesenchymal cell population signaling and intercellular communication that directs major tissue restructuring and the shift out of a dormant fasted state by activating both embryonic developmental and wound healing pathways. We also identify distinct roles of BEST4+ enterocytes in coordinating key regenerative transitions via NOTCH signaling. Python intestinal regeneration shares key signaling features and molecules with mammalian gastric bypass, indicating that conserved regenerative programs are common to both. Our findings provide different insights into cooperative and conserved regenerative programs and intercellular interactions in vertebrates independent of crypts which have been otherwise obscured in model species where temporal phases of generative growth are limited to embryonic development or recovery from injury. [ABSTRACT FROM AUTHOR]

tRNAs are evolutionarily ancient molecular decoders essential for protein translation. In eukaryotes, tRNAs and other short, noncoding RNAs are transcribed by RNA polymerase (Pol) III, an enzyme that promotes ageing in yeast, worms, and flies. Here, we show that a partial reduction in Pol III activity specifically disrupts tRNA levels. This effect is conserved across worms, flies, and mice, where computational models indicate that it impacts mRNA decoding. In all 3 species, reduced Pol III activity increases proteostatic resilience. In worms, it activates the unfolded protein response (UPR) and direct disruption of tRNA metabolism is sufficient to recapitulate this. In flies, decreasing Pol III's transcriptional initiation on tRNA genes by a loss-of-function in the TFIIIC transcription factor robustly extends lifespan, improves proteostatic resilience and recapitulates the broad-spectrum benefits to late-life health seen following partial Pol III inhibition. We provide evidence that a partial reduction in Pol III activity impacts translation, quantitatively or qualitatively, in both worms and flies, indicating a potential mode of action. Our work demonstrates a conserved and previously unappreciated role of tRNAs in animal ageing. tRNAs are important for protein translation, but whether they influence aging is unknown. In this paper, by manipulating RNA polymerase III-mediated transcription of tRNAs across three animal species, the authors demonstrate previously unappreciated roles of tRNA in proteostasis, old-age health and animal longevity. [ABSTRACT FROM AUTHOR]

Morphogens play a critical role in coordinating stress adaptation and aging across tissues, yet their involvement in neuronal mitochondrial stress responses and systemic effects remains unclear. In this study, we reveal that the transforming growth factor beta (TGF-β) DAF-7 is pivotal in mediating the intestinal mitochondrial unfolded protein response (UPRmt) in Caenorhabditis elegans under neuronal mitochondrial stress. Two ASI sensory neurons produce DAF-7, which targets DAF-1/TGF-β receptors on RIM interneurons to orchestrate a systemic UPRmt response. Remarkably, inducing mitochondrial stress specifically in ASI neurons activates intestinal UPRmt, extends lifespan, enhances pathogen resistance, and reduces both brood size and body fat levels. Furthermore, dopamine positively regulates this UPRmt activation, while GABA acts as a systemic suppressor. This study uncovers the intricate mechanisms of systemic mitochondrial stress regulation, emphasizing the vital role of TGF-β in metabolic adaptations that are crucial for organismal fitness and aging during neuronal mitochondrial stress. Neurons play a vital role in inter-tissue communications. Here, authors show the ASI-RIM neuronal axis regulates mitochondrial stress across tissues via TGF-β signaling in C. elegans, linking neuronal mito-stress to several physiological changes. [ABSTRACT FROM AUTHOR]

Conserved signaling cascades monitor protein-folding homeostasis to ensure proper cellular function. One of the evolutionary conserved key players is IRE1, which maintains endoplasmic reticulum (ER) homeostasis through the unfolded protein response (UPR). Upon accumulation of misfolded proteins in the ER, IRE1 forms clusters on the ER membrane to initiate UPR signaling. What regulates IRE1 cluster formation is not fully understood. Here, we show that the ER lumenal domain (LD) of human IRE1α forms biomolecular condensates in vitro. IRE1α LD condensates were stabilized both by binding to unfolded polypeptides as well as by tethering to model membranes, suggesting their role in assembling IRE1α into signaling-competent stable clusters. Molecular dynamics simulations indicated that weak multivalent interactions drive IRE1α LD clustering. Mutagenesis experiments identified disordered regions in IRE1α LD to control its clustering in vitro and in cells. Importantly, dysregulated clustering of IRE1α mutants led to defects in IRE1α signaling. Our results revealed that disordered regions in IRE1α LD control its clustering and suggest their role as a common strategy in regulating protein assembly on membranes. Synopsis: Proteotoxic stress causes the endoplasmic reticulum (ER) stress sensor IRE1α to cluster, but the mechanistic basis of its cluster formation remains poorly understood. This study reveals that disordered regions within the protein's ER lumenal domain (LD) mediate its assembly into signaling clusters. IRE1α LD forms biomolecular condensates in solution. Disordered regions within IRE1α LD drive the formation of condensates through weak multivalent interactions. Binding of unfolded polypeptide ligands and membrane-tethering leads to the formation of stable IRE1α LD clusters. The integrity of the disordered regions in its lumenal domain is critical for IRE1α clustering and unfolded protein response (UPR) signaling in cells. The disordered regions of the ER luminal domain of IRE1α form condensates in vitro and control IRE1α clustering and stress signalling in cells. [ABSTRACT FROM AUTHOR]

Prion disease is a fatal and infectious neurodegenerative disorder caused by the trans-conformation conversion of PRNP/PrPC to PRNP/PrPSc. Accumulated PRNP/PrPSc-induced ER stress causes chronic unfolded protein response (UPR) activation, which is one of the fundamental steps in prion disease progression. However, the role of various ER-resident proteins in prion-induced ER stress is elusive. This study demonstrated that ARL6IP5 is compensatory upregulated in response to chronically activated UPR in the cellular prion disease model (RML-ScN2a). Furthermore, overexpression of ARL6IP5 overcomes ER stress by lowering the expression of chronically activated UPR pathway proteins. We discovered that ARL6IP5 induces reticulophagy to reduce the PRNP/PrPSc burden by releasing ER stress. Conversely, the knockdown of ARL6IP5 leads to inefficient macroautophagic/autophagic flux and elevated PRNP/PrPSc burden. Our study also uncovered that ARL6IP5-induced reticulophagy depends on Ca2+-mediated AMPK activation and can induce 3 MA-inhibited autophagic flux. The detailed mechanistic study revealed that ARL6IP5-induced reticulophagy involves interaction with soluble reticulophagy receptor CALCOCO1 and lysosomal marker LAMP1, leading to degradation in lysosomes. Here, we delineate the role of ARL6IP5 as a novel ER stress regulator and reticulophagy inducer that can effectively reduce the misfolded PRNP/PrPSc burden. Our research opens up a new avenue of selective autophagy in prion disease and represents a potential therapeutic target.Abbreviations: ARL6IP5: ADP ribosylation factor-like GTPase 6 interacting protein 5; AMPK: adenosine 5’-monophosphate (AMP)-activated protein kinase; CALCOCO1: calcium binding and coiled-coil domain 1; CQ: chloroquine; DAPI: 4’6-diamino-2-phenylindole; ER: endoplasmic reticulum; ERPHS: reticulophagy/ER-phagy sites; KD: knockdown; KD-CON: knockdown control; LAMP1: lysosomal-associated membrane protein 1; MAP1LC3/LC3, microtubule-associated protein 1 light chain 3; MTOR: mechanistic target of rapamycin kinase; MβCD: methyl beta cyclodextrin; 3 MA: 3-methyladenine; OE: overexpression; OE-CON: empty vector control; PrDs: prion diseases; PRNP/PrPC: cellular prion protein (Kanno blood group); PRNP/PrPSc: infectious scrapie misfolded PRNP; Tm: tunicamycin; UPR: unfolded protein response; UPS: ubiquitin-proteasome system. [ABSTRACT FROM AUTHOR]

Unfolded protein response (UPR) is a central stress response pathway that is hijacked by tumor cells for their survival. Here, we find that IRE1α signaling, one of the canonical UPR arms, is increased in prostate cancer (PCa) patient tumors. Genetic or small molecule inhibition of IRE1α in syngeneic mouse PCa models and an orthotopic model decreases tumor growth. IRE1α ablation in cancer cells potentiates interferon responses and activates immune system related pathways in the tumor microenvironment (TME). Single-cell RNA-sequencing analysis reveals that targeting IRE1α in cancer cells reduces tumor-associated macrophage abundance. Consistently, the small molecule IRE1α inhibitor MKC8866, currently in clinical trials, reprograms the TME and enhances anti-PD-1 therapy. Our findings show that IRE1α signaling not only promotes cancer cell growth and survival but also interferes with anti-tumor immunity in the TME. Thus, targeting IRE1α can be a promising approach for improving anti-PD-1 immunotherapy in PCa. The IRE1α-XBP1s pathway has been implicated in the regulation of anti-tumor immunity. Here the authors show that IRE1α is increased in prostate cancer (PCa) and that its inhibition reprograms the tumor microenvironment, promoting anti-tumor immune responses in PCa preclinical models. [ABSTRACT FROM AUTHOR]

Non-alcoholic fatty liver disease (NAFLD), a spectrum of liver conditions characterized by fat accumulation without excessive alcohol consumption, represents a significant global health burden. The intricate molecular landscape underlying NAFLD pathogenesis involves lipid handling, inflammation, oxidative stress, and mitochondrial dysfunction, with endoplasmic reticulum (ER) stress emerging as a key contributor. ER stress triggers the unfolded protein response (UPR), impacting hepatic steatosis in NAFLD and contributing to inflammation, fibrosis, and progression to NASH and eventually hepatocellular carcinoma (HCC). Heat shock proteins (HSPs), including small HSPs such as HSP20 and HSP27, HSP60, HSP70, GRP78, and HSP90, are integral to cellular stress responses. They aid in protein folding, prevent aggregation, and facilitate degradation, thus mitigating cellular damage under stress conditions. In NAFLD, aberrant HSP expression and function contribute to disease pathogenesis. Understanding the specific roles of HSP subtypes in NAFLD offers insights into potential therapeutic interventions. This review discusses the involvement of HSPs in NAFLD pathophysiology and highlights their therapeutic potential. By elucidating the molecular mechanisms underlying HSP-mediated protection in NAFLD, this article aims to pave the way for the development of targeted therapies for this prevalent liver disorder. [ABSTRACT FROM AUTHOR]

Common single-nucleotide variants (SNVs) of eukaryotic translation initiation factor 2 alpha kinase 3 (EIF2AK3) slightly increase the risk of disorders in the periphery and the central nervous system. EIF2AK3 encodes protein kinase RNA-like endoplasmic reticulum kinase (PERK), a key regulator of ER stress. Three exonic EIF2AK3 SNVs form the PERK-B haplotype, which is present in 28% of the global population. Importantly, the precise impact of these SNVs on PERK activity remains elusive. In this study, we demonstrate that PERK-B SNVs do not alter PERK expression or basal activity in vitro and in the novel triple knock-in mice expressing the exonic PERK-B SNVs in vivo. However, the kinase activity of PERK-B protein is higher than that of PERK-A in a cell-free assay and in mouse liver homogenates. Pancreatic tissue in PERK-B/B mice also exhibit increased susceptibility to apoptosis under acute ER stress. Monocyte-derived macrophages from PERK-B/B mice exhibit higher PERK activity than those from PERK-A/A mice, albeit with minimal functional consequences at acute timepoints. The subtle PERK-B-driven effects observed in liver and pancreas during acute stress implicate PERK as a contributor to disease susceptibility. The novel PERK-B mouse model provides valuable insights into ER stress-induced PERK activity, aiding the understanding of the genetic basis of disorders associated with ER stress. [ABSTRACT FROM AUTHOR]

All viruses are obligate intracellular parasites that use host machinery to synthesize viral proteins. In infected eukaryotes, viral secreted and transmembrane proteins are synthesized at the endoplasmic reticulum (ER). Many viruses refashion ER membranes into bespoke factories where viral products accumulate while evading host pattern recognition receptors. ER processes are tightly regulated to maintain cellular homeostasis, so viruses must either conform to ER regulatory mechanisms or subvert them to ensure efficient viral replication. Reticulophagy is a catabolic process that directs lysosomal degradation of ER components. There is accumulating evidence that reticulophagy serves as a form of antiviral defense; we call this defense “xERophagy” to acknowledge its relationship to xenophagy, the catabolic degradation of microorganisms by macroautophagy/autophagy. In turn, viruses can subvert reticulophagy to suppress host antiviral responses and support efficient viral replication. Here, we review the evidence for functional interplay between viruses and the host reticulophagy machinery.Abbreviations: AMFR: autocrine motility factor receptor; ARF4: ADP-ribosylation factor 4; ARL6IP1: ADP-ribosylation factor-like 6 interacting protein 1; ATL3: atlastin GTPase 3; ATF4: activating transcription factor 4; ATF6: activating transcription factor 6; BPIFB3: BPI fold containing family B, member 3; CALCOCO1: calcium binding and coiled coil domain 1; CAMK2B: calcium/calmodulin-dependent protein kinase II, beta; CANX: calnexin; CDV: canine distemper virus; CCPG1: cell cycle progression 1; CDK5RAP3/C53: CDK5 regulatory subunit associated protein 3; CIR: cargo-interacting region; CoV: coronavirus; CSNK2/CK2: casein kinase 2; CVB3: coxsackievirus B3; DAPK1: death associated protein kinase 1; DENV: dengue virus; DMV: double-membrane vesicles; EBOV: Ebola virus; EBV: Epstein-Barr Virus; EIF2AK3/PERK: eukaryotic translation initiation factor 2 alpha kinase 3; EMCV: encephalomyocarditis virus; EMV: extracellular microvesicle; ER: endoplasmic reticulum; ERAD: ER-associated degradation; ERN1/IRE1: endoplasmic reticulum to nucleus signalling 1; EV: extracellular vesicle; EV71: enterovirus 71; FIR: RB1CC1/FIP200-interacting region; FMDV: foot-and-mouth disease virus; HCMV: human cytomegalovirus; HCV: hepatitis C virus; HMGB1: high mobility group box 1; HSPA5/BiP: heat shock protein 5; IFN: interferon; IFNG/IFN-γ: interferon gamma; KSHV: Kaposi’s sarcoma-associated herpesvirus; LIR: MAP1LC3/LC3-interacting region; LNP: lunapark, ER junction formation factor; MAP1LC3: microtubule-associated protein 1 light chain 3; MAP3K5/ASK1: mitogen-activated protein kinase kinase kinase 5; MAPK/JNK: mitogen-activated protein kinase; MeV: measles virus; MHV: murine hepatitis virus; NS: non-structural; PDIA3: protein disulfide isomerase associated 3; PRR: pattern recognition receptor; PRRSV: porcine reproductive and respiratory syndrome virus; RB1CC1/FIP200: RB1-inducible coiled-coil 1; RETREG1/FAM134B: reticulophagy regulator 1; RHD: reticulon homology domain; RTN3: reticulon 3; RTN3L: reticulon 3 long; sAIMs: shuffled Atg8-interacting motifs; SARS-CoV: severe acute respiratory syndrome coronavirus; SINV: Sindbis virus; STING1: stimulator of interferon response cGAMP interactor 1; SVV: Seneca Valley virus; SV40: simian virus 40; TEX264: testis expressed gene 264 ER-phagy receptor; TFEB: transcription factor EB; TRAF2: TNF receptor-associated factor 2; UIM: ubiquitin-interacting motif; UFM1: ubiquitin-fold modifier 1; UPR: unfolded protein response; VAPA: vesicle-associated membrane protein, associated protein A; VAPB: vesicle-associated membrane protein, associated protein B and C; VZV: varicella zoster virus; WNV: West Nile virus; XBP1: X-box binding protein 1; XBP1s: XBP1 spliced; xERophagy: xenophagy involving reticulophagy; ZIKV: Zika virus [ABSTRACT FROM AUTHOR]

The cytokine tumor necrosis factor (TNF) plays important roles in limiting infection but is also linked to sepsis. The mechanisms underlying these paradoxical roles are unclear. Here, we show that TNF limits the antimicrobial activity of Paneth cells (PCs), causing bacterial translocation from the gut to various organs. This TNF-induced lethality does not occur in mice with a PC-specific deletion in the TNF receptor, P55. In PCs, TNF stimulates the IFN pathway and ablates the steady-state unfolded protein response (UPR), effects not observed in mice lacking P55 or IFNAR1. TNF triggers the transcriptional downregulation of IRE1 key genes Ern1 and Ern2 , which are key mediators of the UPR. This UPR deficiency causes a significant reduction in antimicrobial peptide production and PC antimicrobial activity, causing bacterial translocation to organs and subsequent polymicrobial sepsis, organ failure, and death. This study highlights the roles of PCs in bacterial control and therapeutic targets for sepsis. [Display omitted] • PC-specific TNFR1 (P55) mutant mice are protected against lethal TNF effects • PC-P55 signaling triggers an interferon signature and UPR failure in these cells • This UPR failure in PCs is IFNAR1 and P55 dependent but microbiome independent • PC-P55 UPR failure reduces AMP activity, causing bacterial escape and sepsis Wallaeys et al. studied Paneth cells (PCs) during TNF-induced inflammation. TNF signaling through its P55 receptor on PCs triggers an interferon signature and disrupts the unfolded protein response, causing reduced production of antimicrobial peptides and decreased antimicrobial activity. This PC dysfunction causes gut bacterial translocation and sepsis. [ABSTRACT FROM AUTHOR]

Fungal HacA/Hac1 transcription factors play a crucial role in regulating the unfolded protein response (UPR). The UPR helps cells to maintain endoplasmic reticulum (ER) protein homeostasis, which is critical for growth, development, and virulence. The Aspergillus flavus hacA gene encodes a domain rich in basic and acidic amino acids (Bsc) and a basic leucine zipper (bZip) domain, and features a non-conventional intron (Nt20). In this study, CRISPR/Cas9 was utilized to dissect the Bsc-coding, bZip-coding, and Nt20 sequences to elucidate the relationship between genotype and phenotype. In the Bsc and bZip experimental sets, all observed mutations in both coding sequences were in frame, suggesting that out-of-frame mutations are lethal. The survival rate of transformants in the Nt20 experiment set was low, at approximately 7%. Mutations in the intron primarily consisted of out-of-frame insertions and deletions. In addition to the wild-type-like conidial morphology, the mutants exhibited varied colony morphologies, including sclerotial, mixed (conidial and sclerotial), and mycelial morphologies. An ER stress test using dithiothreitol revealed that the sclerotial and mycelial mutants were much more sensitive than the conidial mutants. Additionally, the mycelial mutants were unable to produce aflatoxin but still produced aspergillic acid and kojic acid. RNAi experiments targeting the region encompassing Bsc and bZip indicated that transformant survival rates generally decreased, with a small number of transformants displaying phenotypic changes. Defects in the hacA gene at the DNA and transcript levels affected the survival, growth, and development of A. flavus. Thus, this gene may serve as a promising target for future host-induced gene-silencing strategies aimed at controlling infection and reducing aflatoxin contamination in crops. [ABSTRACT FROM AUTHOR]

Simple Summary: This study investigates how calcium oxalate crystals interact with renal tubular epithelial cells and contribute to kidney stone formation. By using transcriptome sequencing, the study identifies 629 differentially expressed genes when renal tubular epithelial cells are exposed to calcium oxalate crystals, with a significant number of these genes linked to endoplasmic reticulum stress and unfolded protein response. The study also highlights the potential crucial role of two specific genes, CHAC1 and FGF21, which were found to be significantly up-regulated in both cell models and kidney tissues from patients with calcium oxalate stones. The study suggests that targeting endoplasmic reticulum stress-related molecules and pathways could offer new therapeutic strategies to prevent stone formation and recurrence. Our study aimed to elucidate the mechanisms behind the interaction between calcium oxalate (CaOx) crystals and renal tubular epithelial cells through transcriptome sequencing analysis. HK-2 cells were stimulated with or without CaOx monohydrate crystals and subjected to RNA-seq to assess the effects of CaOx crystals on gene expression changes, key pathways, and molecular players during this interaction. A total of 629 differentially expressed genes (DEGs) were identified between the control group and experimental group, with 491 genes up-regulated and 138 down-regulated. Functional enrichment analysis indicated that the DEGs were significantly associated with endoplasmic reticulum stress (ERS) and unfolded protein response. To validate our findings, we compared our results with the public dataset GSE73680 and confirmed the increased expression of two ERS-related DEGs, CHAC1 and FGF21, in renal papillary tissues from patients with CaOx stones. Collectively, these findings suggest that ERS plays a crucial role in the crystal–cell interaction and highlight the potential for developing therapeutic strategies aimed at reducing CaOx stone formation by targeting ERS-related molecules and pathways. [ABSTRACT FROM AUTHOR]

In eukaryotic cells, the endoplasmic reticulum is particularly important in post-translational modification of proteins before they are released extracellularly or sent to another endomembrane system. The correct three-dimensional folding of most proteins occurs in the ER lumen, which has an oxidative environment that is essential for the formation of disulfide bridges, which are important in maintaining protein structure. The ER is a versatile organelle that ensures the correct structure of proteins and is essential in the synthesis of lipids and sterols, in addition to offering support in the maintenance of intracellular calcium. Consequently, the cells needed to respond to demands caused by physiological conditions and pathological disturbances in the organelle homeostasis, leading to proper functioning of the cell or even programmed cell death. Disturbances to the ER function trigger a response to the accumulation of unfolded or misfolded proteins, known as the unfolded protein response. Such disturbances include abiotic stress, pharmacological agents, and intracellular pathogens, such as viruses. When misfolded proteins accumulate in the ER, they can undergo ubiquitination and proteasomal degradation through components of the ER-associated degradation system. Once a prolonged activity of the UPR pathway occurs, indicating that homeostasis cannot be reestablished, components of this pathway induce cell death by apoptosis. Here, we discuss how viruses have evolved ways to counteract UPR responses to maximize replication. This evolutionary viral ability is important to understand cell pathology and should be taken into account when designing therapeutic interventions and vaccines. [ABSTRACT FROM AUTHOR]

The human GBA1 gene encodes lysosomal acid β-glucocerebrosidase, whose activity is deficient in Gaucher disease (GD). In Drosophila, there are two GBA1 orthologs, Gba1a and Gba1b, and Gba1b is the bona fide GCase encoding gene. Several fly lines with different deletions in the Gba1b were studied in the past. However, since most GD-associated GBA1 mutations are point mutations, we created missense mutations homologous to the two most common GD mutations: the mild N370S mutation (D415S in Drosophila) and the severe L444P mutation (L494P in Drosophila), using the CRISPR-Cas9 technology. Flies homozygous for the D415S mutation (dubbed D370S hereafter) presented low GCase activity and substrate accumulation, which led to lysosomal defects, activation of the Unfolded Protein Response (UPR), inflammation/neuroinflammation, and neurodegeneration along with earlier death compared to control flies. Surprisingly, the L494P (called L444P hereafter) flies presented higher GCase activity with fewer lysosomal defects and milder disease in comparison to that presented by the D370S homozygous flies. Treatment with ambroxol had a limited effect on all homozygous fly lines tested. Overall, our results underscore the differences between the fly and human GCase enzymes, as evidenced by the distinct phenotypic outcomes of mutations in flies compared to those observed in human GD patients. [ABSTRACT FROM AUTHOR]

As a component of circulating lipoproteins, APOE binds to cell surface receptors mediating lipoprotein metabolism and cholesterol transport. A growing body of evidence, including the identification of a broad variety of cellular proteins interacting with APOE, suggests additional independent functions. Investigating cellular localization and protein–protein interactions in cultured human hepatocytes, we aimed to contribute to the elucidation of hitherto unnoted cellular functions of APOE. We observed a strong accumulation of APOE in MAMs, equally evident for the two major isoforms APOE3 and APOE4. Using mass spectrometry proteome analyses, novel and previously noted APOE interactors were identified, including the mitochondrial proteins TOMM40, LONP1 and VDAC1. All three interactors were present in MAM fractions, which we think initially facilitates interactions with APOE. LONP1 is a protease with chaperone activity, which migrated to MAMs in response to ER stress, displaying a reinforced interaction with APOE. We therefore hypothesize that APOE may help in the unfolded protein response (UPR) by acting as a co-chaperone in cooperation with LONP1 at the interface of mitochondria and ER membranes. The interaction of APOE with the integral proteins TOMM40 and VDAC1 may point to the formation of bridging complexes connecting mitochondria with other organelles. [ABSTRACT FROM AUTHOR]

2-deoxy-D-glucose (2DG) is a glycolysis and protein N-glycosylation inhibitor with promising anti-tumor and immunomodulatory effects. However, 2DG can also suppress T cell function, including IFN-γ secretion. Few human T cell studies have studied low-dose 2DG, which can increase IFN-γ in a Jurkat clone. We therefore investigated 2DG's effect on IFN-γ in activated human T cells from PBMCs, with 2DG treatment commenced either concurrently with activation or 48 h after activation. Concurrent 2DG treatment decreased IFN-γ secretion in a dose-dependent manner. However, 2DG treatment of pre-activated T cells had a hormetic effect on IFN-γ, with 0.15–0.6 mM 2DG (achievable in vivo) increasing and >2.4 mM 2DG reducing its secretion. In contrast, IL-2 levels declined monotonously with increasing 2DG concentration. Lower 2DG concentrations reduced PD-1 and increased CD69 expression regardless of treatment timing. The absence of increased T-bet or Eomes expression or IFNG transcription suggests another downstream mechanism. 2DG dose-dependently induced the unfolded protein response, suggesting a possible role in increased IFN-γ secretion, possibly by increasing the ER folding capacity for IFN-γ via increased chaperone expression. Overall, low-dose, short-term 2DG exposure could potentially improve the T cell anti-tumor response. [ABSTRACT FROM AUTHOR]

Cerebral dopamine neurotrophic factor (CDNF) is an unconventional neurotrophic factor because it does not bind to a known specific receptor on the plasma membrane and functions primarily as an unfolded protein response (UPR) regulator in the endoplasmic reticulum. Data on the effects of CDNF on nonmotor behavior and monoamine metabolism are limited. Here, we performed the intracerebroventricular injection of a recombinant CDNF protein at doses of 3, 10, and 30 μg in C57BL/6 mice. No adverse effects of the CDNF injection on feed and water consumption or locomotor activity were observed for 3 days afterwards. Decreases in body weight and sleep duration were transient. CDNF-treated animals demonstrated improved performance on the operant learning task and a substantial decrease in anxiety and behavioral despair. CDNF in all the doses enhanced serotonin (5-HT) turnover in the murine frontal cortex, hippocampus, and midbrain. This alteration was accompanied by changes in the mRNA levels of the 5-HT1A and 5-HT7 receptors and in monoamine oxidase A mRNA and protein levels. We found that CDNF dramatically increased c-Fos mRNA levels in all investigated brain areas but elevated the phosphorylated-c-Fos level only in the midbrain. Similarly, enhanced CREB phosphorylation was found in the midbrain in experimental animals. Additionally, the upregulation of a spliced transcript of XBP1 (UPR regulator) was detected in the midbrain and frontal cortex. Thus, we can hypothesize that exogenous CDNF modulates the UPR pathway and overall neuronal activation and enhances 5-HT turnover, thereby affecting learning and emotion-related behavior. [ABSTRACT FROM AUTHOR]

Osteoblasts and osteoclasts collaborate in bone metabolism, facilitating bone development, maintaining normal bone density and strength, and aiding in the repair of pathological damage. Endoplasmic reticulum stress (ERS) can disrupt the intracellular equilibrium between osteoclast and osteoblast, resulting in dysfunctional bone metabolism. The inositol‐requiring enzyme‐1α (IRE1α) pathway—the most conservative unfolded protein response pathway activated by ERS—is crucial in regulating cell metabolism. This involvement encompasses functions such as inflammation, autophagy, and apoptosis. Many studies have highlighted the potential roles of the IRE1α pathway in osteoblasts, chondrocytes, and osteoclasts and its implication in certain bone‐related diseases. These findings suggest that it may serve as a mediator for bone metabolism. However, relevant reviews on the role of the IRE1α pathway in bone metabolism remain unavailable. Therefore, this review aims to explore recent research that elucidated the intricate roles of the IRE1α pathway in bone metabolism, specifically in osteogenesis, chondrogenesis, osteoclastogenesis, and osteo‐immunology. The findings may provide novel insights into regulating bone metabolism and treating bone‐related diseases. [ABSTRACT FROM AUTHOR]

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. [ABSTRACT FROM AUTHOR]

Endoplasmic reticulum stress triggers the unfolded protein response (UPR) to promote cell survival or apoptosis. Transient endoplasmic reticulum stress activation has been reported to trigger megakaryocyte production, and UPR activation has been reported as a feature of megakaryocytic cancers. However, the role of UPR signaling in megakaryocyte biology is not fully understood. We studied the involvement of UPR in human megakaryocytic differentiation using PMA (phorbol 12‐myristate 13‐acetate)‐induced maturation of megakaryoblastic cell lines and thrombopoietin‐induced differentiation of human peripheral blood‐derived progenitors. Our results demonstrate that an adaptive UPR is a feature of megakaryocytic differentiation and that this response is not associated with ER stress‐induced apoptosis. Differentiation did not alter the response to the canonical endoplasmic reticulum stressors DTT or thapsigargin. However, thapsigargin, but not DTT, inhibited differentiation, consistent with the involvement of Ca2+ signaling in megakaryocyte differentiation. [ABSTRACT FROM AUTHOR]

Activating transcription factor 4 (ATF4) plays a central role in the integrated stress response (ISR) and one overlapping branch of the unfolded protein response (UPR). We recently reported that the splicing inhibitor isoginkgetin (IGG) induced ATF4 protein along with several known ATF4-regulated transcripts in a response that resembled the ISR and UPR. However, the contribution of ATF4-dependent and -independent transcriptional responses to IGG exposure was not known. Here we used RNA-sequencing in HCT116 colon cancer cells and an isogenic subline lacking ATF4 to investigate the contribution of ATF4 to IGG-induced changes in gene expression. Approximately 85% of the IGG-responsive DEGs in HCT116 cells were also differentially expressed in response to the ER stressor thapsigargin (Tg) and these were enriched for genes associated with the UPR and ISR. Most of these were positively regulated by IGG with impaired responses in the ATF4-deficient cells. Nonetheless, there were DEGs that responded similarly in both cell lines. The ATF4-independent IGG-induced DEGs included several metal responsive transcripts encoding metallothionines and a zinc transporter. Taken together, the predominant IGG response was ATF4-dependent in these cells and resembled the UPR and ISR while a second less prominent response involved the ATF4-independent regulation of metal responsive mRNAs. [ABSTRACT FROM AUTHOR]

Splicing and endoplasmic reticulum (ER)-proteostasis are two key processes that ultimately regulate the functional proteins that are produced by a cell. However, the extent to which these processes interact remains poorly understood. Here, we identify SNRPB and other components of the Sm-ring, as targets of the unfolded protein response and novel regulators of export from the ER. Mechanistically, The Sm-ring regulates the splicing of components of the ER export machinery, including Sec16A, a component of ER exit sites. Loss of function of SNRPB is causally linked to cerebro-costo-mandibular syndrome (CCMS), a genetic disease characterized by bone defects. We show that heterozygous deletion of SNRPB in mice resulted in bone defects reminiscent of CCMS and that knockdown of SNRPB delays the trafficking of type-I collagen. Silencing SNRPB inhibited osteogenesis in vitro, which could be rescued by overexpression of Sec16A. This rescue indicates that the role of SNRPB in osteogenesis is linked to its effects on ER-export. Finally, we show that SNRPB is a target for the unfolded protein response, which supports a mechanistic link between the spliceosome and ER-proteostasis. Our work highlights components of the Sm-ring as a novel node in the proteostasis network, shedding light on CCMS pathophysiology. Synopsis: RNA splicing and endoplasmic reticulum (ER) proteostasis are key to the overall control of mature protein production rates; however, the interaction between these processes is not well understood. In the following work, the spliceosome is shown to be a target of the unfolded protein response and a regulator of type-I collagen export. Silencing SNRPB and other spliceosome components alters ER export by regulating the splicing of Sec16A Loss of SNRPB results in type-I collagen trafficking defects in cell culture and osteogenesis defects in vivo SNRPB is regulated by the unfolded protein response (UPR) Our work contributes towards a better understanding of cerebro-costo-mandibular syndrome, a disease that is caused by loss of SNRPB Loss of SM-proteins alters the splicing of ER export factors which causes defects in ER-proteostasis and protein secretion. [ABSTRACT FROM AUTHOR]

Background: Tobacco smoking is the leading cause of preventable death and disease worldwide, with over 8 million annual deaths attributed to cigarette smoking. This study investigates the impact of cigarette smoke and heated tobacco products (HTPs) on microglial function, focusing on toxicological profiles, inflammatory responses, and oxidative stress using ISO standard and clinically relevant conditions of exposure. Methods: We assessed cell viability, reactive oxygen species (ROS) production, lipid peroxidation, mitochondrial function, unfolded protein response, and inflammation in human microglial cells (HMC3) exposed to cigarette smoke, HTP aerosol or nicotine. Results: Our findings show that cigarette smoke significantly reduces microglial viability, increases ROS formation, induces lipid peroxidation, and reduces intracellular glutathione levels. Cigarette smoke also alters the expression of genes involved in mitochondrial dynamics and biogenesis, leading to mitochondrial dysfunction. Additionally, cigarette smoke impairs the unfolded protein response, activates the NF-κB pathway, and induces a pro-inflammatory state characterized by increased TNF and IL-18 expression. Furthermore, cigarette smoke causes DNA damage and decreases the expression of the aging marker Klotho β. In contrast, HTP, exhibited a lesser degree of microglial toxicity, with reduced ROS production, lipid peroxidation, and mitochondrial dysfunction compared to conventional cigarettes. Conclusion: These results highlight the differential toxicological profile of cigarette smoke and HTP on microglial cells, suggesting a potential harm reduction strategy for neurodegenerative disease for smokers unwilling or unable to quit. [ABSTRACT FROM AUTHOR]

A study published in the journal Translational Neurodegeneration examines the role of the integrated stress response (ISR) in amyotrophic lateral sclerosis (ALS). The researchers specifically investigate the impact of inhibiting the ISR kinase GCN2 on the clustering and toxicity of mutant SOD1, a protein associated with ALS. The study finds that inhibiting GCN2 delays disease progression in a mouse model of ALS by reducing mutant SOD1 clustering. These findings suggest that targeting the ISR pathway, particularly GCN2, may hold promise for ALS treatment. [Extracted from the article]

Background: Alpha-1 antitrypsin deficiency (AATD) is an inherited disease, the common variant caused by a Pi*Z mutation in the SERPINA1 gene. Pi*Z AAT increases the risk of pulmonary emphysema and liver disease. Berberine (BBR) is a nature dietary supplement and herbal remedy. Emerging evidence revealed that BBR has remarkable liver-protective properties against various liver diseases. In the present study, we investigated the therapeutic effects and toxicities of BBR in Pi*Z hepatocytes and Pi*Z transgenic mice. Methods: Huh7.5 and Huh7.5Z (which carries the Pi*Z mutation) cells were treated with different concentrations of BBR for 48 hours. MTT was performed for cell viability assay. Intracellular AAT levels were evaluated by western blot. In vivo studies were carried out in wild type, native phenotype AAT (Pi*M), and Pi*Z AAT transgenic mice. Mice were treated with 50 mg/kg/day of BBR or solvent only by oral administration for 30 days. Western blot and liver histopathological examinations were performed to evaluate therapeutic benefits and liver toxicity of BBR. Results: BBR reduced intracellular AAT levels in Huh7.5Z cells, meanwhile, no Pi*Z-specific toxicity was observed. However, BBR did not reduce liver AAT load but significantly potentiated liver inflammation and fibrosis accompanying the activation of unfolded protein response and mTOR in Pi*Z mice, but not in wild type and Pi*M mice. Conclusions: BBR exacerbated liver inflammation and fibrosis specifically in Pi*Z mice. This adverse effect may be associated with the activation of unfolded protein response and mTOR. This study implicates that BBR should be avoided by AATD patients. [ABSTRACT FROM AUTHOR]

Background: Myocardial infarction (MI) significantly contributes to the global mortality rate, often leading to heart failure (HF) due to left ventricular remodeling. Key factors in the pathomechanism of HF include nitrosative/oxidative stress, inflammation, and endoplasmic reticulum (ER) stress. Furthermore, while a high-fat diet (HFD) is known to exacerbate post-MI cardiac remodeling, its impact on these critical factors in the context of HF is not as well understood. Aims: This study aimed to assess the impact of post-MI HF and HFD on inflammation, nitro-oxidative stress, ER stress, and unfolded protein response (UPR). Methods: The study was performed on fragments of the left ventricle harvested from 30 male adult Sprague Dawley rats, which were divided into four groups based on diet (normal-fat vs. high-fat) and surgical procedure (sham operation vs. coronary artery ligation to induce MI). We assessed body weight, NT-proBNP levels, protein levels related to nitrosative/oxidative stress, ER stress, UPR, apoptosis, and nitric oxide synthases, through Western Blot and ELISA. Results: HFD and MI significantly influenced body weight and NT-proBNP concentrations. HFD elevated 3-nitrotyrosine and myeloperoxidase levels and altered nitric oxide synthase levels. HFD and MI significantly affected ER stress markers and activated or inhibited UPR pathways. Conclusions: The study demonstrates significant impacts of post-MI HF and dietary fat content on cardiac function and stress markers in a rat model. The interaction between HFD and MI on UPR activation suggests the importance of dietary management in post-MI recovery and HF prevention. [ABSTRACT FROM AUTHOR]

Lsm14b (LSM family member 14b) is a messenger ribonucleoprotein (mRNP) and a widely present component in eukaryotes. Lsm14b participates in oocyte development by regulating mRNA translation, however, the specific translational regulatory mechanisms remain unclear. Here, we explore the function of Lsm14b during early oocyte development and identify specific translational regulatory mechanisms. We established female-infertile lsm14b mutant zebrafish using CRISPR/Cas9. Histological examination showed that the oocyte development in the mutant zebrafish was arrested at the primary growth (PG) stage. The gene set enrichment analysis (GSEA) analysis of the transcriptome revealed that signaling pathways associated with mRNA translation suppression and mRNA poly(A) tail shortening were significantly downregulation in the mutant. The poly(A) tail length (PAT) assay confirmed the lengthen of mRNA poly(A) tail of the oocyte development-related genes zar1 and figla in the mutant. Further studies have suggested that the loss of Lsm14b triggers the unfolded protein response (UPR), which is related to abnormal translation inhibition. Our results demonstrate that Lsm14b assists in maintaining the translation-inhibited state of mRNA by regulating the length of the mRNA poly(A) tail in zebrafish early oocytes, which ensures that the mRNA synthesized and stored during the growth stage of oocytes, is necessary for the normal growth and development of oocytes. [ABSTRACT FROM AUTHOR]

During homeostasis, the endoplasmic reticulum (ER) maintains productive transmembrane and secretory protein folding that is vital for proper cellular function. The ER-resident HSP70 chaperone, binding immunoglobulin protein (BiP), plays a pivotal role in sensing ER stress to activate the unfolded protein response (UPR). BiP function is regulated by the bifunctional enzyme filamentation induced by cyclic-AMP domain protein (FicD) that mediates AMPylation and deAMPylation of BiP in response to changes in ER stress. AMPylated BiP acts as a molecular rheostat to regulate UPR signaling, yet little is known about the molecular consequences of FicD loss. In this study, we investigate the role of FicD in mouse embryonic fibroblast (MEF) response to pharmacologically and metabolically induced ER stress. We find differential BiP AMPylation signatures when comparing robust chemical ER stress inducers to physiological glucose starvation stress and recovery. Wildtype MEFs respond to pharmacological ER stress by down-regulating BiP AMPylation. Conversely, BiP AMPylation in wildtype MEFs increases upon metabolic stress induced by glucose starvation. Deletion of FicD results in widespread gene expression changes under baseline growth conditions. In addition, FicD null MEFs exhibit dampened UPR signaling, altered cell stress recovery response, and unconstrained protein secretion. Taken together, our findings indicate that FicD is important for tampering UPR signaling, stress recovery, and the maintenance of secretory protein homeostasis. [ABSTRACT FROM AUTHOR]

Our study investigates the impact of FGF23 overexpression on SaOS-2 cells to elucidate its role in cellular stress and morphology, contributing to the understanding of skeletal pathologies like X-linked hypophosphatemia (XLH). Using transmission electron microscopy and protein analysis (Western blot), we analyzed the rough endoplasmic reticulum (rER) and mitochondria in SaOS-2 cells with FGF23 overexpression compared to controls. We found significant morphological changes, including enlarged and elongated rER and mitochondria, with increased contact zones, suggesting enhanced interaction and adaptation to elevated protein synthesis and secretion demands. Additionally, we observed higher apoptosis rates of the cells after 24–72 h in vitro and upregulated proteins associated with ER stress and apoptosis, such as CHOP, XBP1 (spliced and unspliced), GRP94, eIF2α, and BAX. These findings indicate a robust activation of the unfolded protein response (UPR) and apoptotic pathways due to FGF23 overexpression. Our results highlight the critical role of ER and mitochondrial interactions in cellular stress responses and provide new insights into the mechanistic link between FGF23 signaling and cellular homeostasis. In conclusion, our study underscores the importance of analyzing UPR-related pathways in the development of therapeutic strategies for skeletal and systemic diseases and contributes to a broader understanding of diseases like XLH. [ABSTRACT FROM AUTHOR]

Simple Summary: Current evidence suggests that protein synthesis dysfunction within the endoplasmic reticulum (ER) of the small intestine in animals with intrauterine growth retardation (IUGR) may be a potential factor contributing to the adverse postnatal outcomes associated with intestinal growth and development. However, little is known about whether the impaired protein-folding capacity of ER triggers the adaptive cellular response known as the unfolded protein response (UPR), as well as the subsequent signaling pathways leading to cellular apoptosis in the intestines of IUGR pigs. Therefore, this study aimed to evaluate the structural integrity of the ER and elucidate the potential signaling cascade of the UPR, which may mitigate the effects of ER stress (ERS) within the intestinal mucosa of IUGR neonates. The results showed that the mitochondrial swelling and ER dilation in the intestinal mucosa, resulting from restricted intrauterine development, occur on the neonatal day. The activation of the IRE1α and PERK pathways within the UPR, in response to ERS, was observed in the intestines of IUGR newborn piglets, leading to apoptosis in intestinal cells mediated by the transcription factor CHOP. Understanding this apoptotic mechanism underlying the structural and functional impairment in the guts of animals with IUGR is crucial for identifying potential preventive strategies. Intrauterine growth retardation (IUGR) in piglets is associated with a high rate of morbidity and mortality after birth due to gut dysfunction, and the underlying mechanisms remain poorly understood. This study selected six pairs of IUGR newborn male piglets and normal birth weight newborn piglets (Large White × Landrace) to investigate differences in intestinal structure and digestive functions, intestinal ERS and apoptosis, intestinal barrier function, and inflammatory response. The results showed that IUGR significantly reduced the jejunal villi height (p < 0.05) and the ratio of villus-height-to-crypt-depth (p = 0.05) in neonatal piglets. Additionally, the microvilli in the jejunum of IUGR neonatal piglets were shorter than those in normal-weight piglets, and swelling of the mitochondria and expansion of the endoplasmic reticulum were observed. IUGR also significantly reduced serum glucose and lactase levels (p < 0.05) while significantly increasing mRNA levels of jejunal IRE1α, EIF2α, CHOP, Bax, Caspase9, Mucin2, Claudin-1, Occludin, ZO-1, Bcl-2, IL-6, and IFN-γ (p < 0.05), as well as GRP78 protein levels in neonatal piglets (p < 0.05). These findings suggest that IUGR impairs intestinal structure and barrier function in newborn piglets by enhancing intestinal inflammatory responses, activating intestinal ERS and the signaling pathways related to the unfolded protein response, thereby inducing ERS-related apoptosis. [ABSTRACT FROM AUTHOR]

Cancer cachexia manifests as whole body wasting, however, the precise mechanisms governing the alterations in skeletal muscle and cardiac anabolism have yet to be fully elucidated. In this study, we explored changes in anabolic processes in both skeletal and cardiac muscles in the Yoshida AH‐130 ascites hepatoma model of cancer cachexia. AH‐130 tumor‐bearing rats experienced significant losses in body weight, skeletal muscle, and heart mass. Skeletal and cardiac muscle loss was associated with decreased ribosomal (r)RNA, and hypophosphorylation of the eukaryotic factor 4E binding protein 1. Endoplasmic reticulum stress was evident by higher activating transcription factor mRNA in skeletal muscle and growth arrest and DNA damage‐inducible protein (GADD)34 mRNA in both skeletal and cardiac muscles. Tumors provoked an increase in tissue expression of interferon‐γ in the heart, while an increase in interleukin‐1β mRNA was apparent in both skeletal and cardiac muscles. We conclude that compromised skeletal muscle and heart mass in the Yoshida AH‐130 ascites hepatoma model involves a marked reduction translational capacity and efficiency. Furthermore, our observations suggest that endoplasmic reticulum stress and tissue production of pro‐inflammatory factors may play a role in the development of skeletal and cardiac muscle wasting. [ABSTRACT FROM AUTHOR]

Background: High-grade serous ovarian carcinoma (HGSOC) is the most common and aggressive subtype of epithelial ovarian carcinoma. It is primarily diagnosed at stage III or IV when the 5-year survival rate ranges between 20% and 40%. Here, we aimed to validate the hypothesis, based on HGSOC cell lines, that proposed the existence of two distinct groups of HGSOC cells with high and low oxidative phosphorylation (OXPHOS) metabolism, respectively, which are associated with their responses to glucose and glutamine withdrawal. Methods: We isolated and cultivated primary cancer cell cultures from HGSOC and nontransformed ovarian fibroblasts from the surrounding ovarium of 45 HGSOC patients. We tested the metabolic flexibility of the primary cells, particularly in response to glucose and glutamine depletion, analyzed and modulated endoplasmic reticulum stress, and searched for indices of the existence of previously reported groups of HGSOC cells with high and low OXPHOS metabolism. Results: The primary HGSOC cells did not form two groups with high and low OXPHOS that responded differently to glucose and glutamine availabilities in the cell culture medium. Instead, they exhibited a continuum of OXPHOS phenotypes. In most tumor cell isolates, the responses to glucose or glutamine withdrawal were mild and surprisingly correlated with those of nontransformed ovarian fibroblasts from the same patients. The growth of tumor-derived cells in the absence of glucose was positively correlated with the lipid trafficking regulator FABP4 and was negatively correlated with the expression levels of HK2 and HK1. The correlations between the expression of electron transport chain (ETC) proteins and the oxygen consumption rates or extracellular acidification rates were weak. ER stress markers were strongly expressed in all the analyzed tumors. ER stress was further potentiated by tunicamycin but not by the recently proposed ER stress inducers based on copper(II)-phenanthroline complexes. ER stress modulation increased autophagy in tumor cell isolates but not in nontransformed ovarian fibroblasts. Conclusions: Analysis of the metabolism of primary HGSOC cells rejects the previously proposed hypothesis that there are distinct groups of HGSOC cells with high and low OXPHOS metabolism that respond differently to glutamine or glucose withdrawal and are characterized by ETC protein levels. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is the most common age‐related neurodegenerative disorder leading to cognitive decline. Excessive cytosolic calcium (Ca2+) accumulation plays a critical role in the pathogenesis of AD since it activates the NOD‐like receptor family, pyrin domain containing 3 (NLRP3), switches the endoplasmic reticulum (ER) unfolded protein response (UPR) toward proapoptotic signaling and promotes Aβ seeding. Herein, a liposomal nanodrug (felodipine@LND) is developed incorporating a calcium channel antagonist felodipine for Alzheimer's disease treatment through a low‐intensity pulse ultrasound (LIPUS) irradiation‐assisted blood brain barrier (BBB)‐crossing drug delivery. The multifunctional felodipine@LND is effectively delivered to diseased brain through applying a LIPUS irradiation to the skull, which resulted in a series of positive effects against AD. Markedly, the nanodrug treatment switched the ER UPR toward antioxidant signaling, prevented the surface translocation of ER calreticulin (CALR) in microglia, and inhibited the NLRP3 activation and Aβ seeding. In addition, it promoted the degradation of damaged mitochondria via mitophagy, thereby inhibiting the neuronal apoptosis. Therefore, the anxiety‐like behavior and cognitive impairment of 5xFAD mice with AD is significantly ameliorated, which manifested the potential of LIPUS – assisted BBB‐crossing delivery of felodipine@LND to serve as a paradigm for AD therapy based on the well‐recognized clinically available felodipine. [ABSTRACT FROM AUTHOR]