Regulator Of Unfolded Protein Response Research Articles

Activation, regulation, and effects of the mitochondrial unfolded protein response in endothelial cells

Abstract Introduction Endoplasmic reticulum stress and the resulting unfolded protein response (UPRER) play significant roles in endothelial dysfunction and atherosclerosis. However, the understanding of the mitochondrial unfolded protein response (UPRMito) in endothelial dysfunction is limited. This study aims to characterize the activation, regulation, and effects of the UPRMito to provide a comprehensive understanding of endothelial UPRMito. Methods and Results RNA sequencing and quantitative proteomic analysis were employed to investigate the impact of stressors on mitochondrial (MitoBlock-6, CDDO) and endoplasmic (Tunicamycin) protein homeostasis on the transcriptome and proteome of Human Coronary Artery Endothelial cells (HCAEC, Fig.1). The analysis identified 12,745 unique transcripts and 7,891 unique proteins. Mitocarta annotation highlighted the regulatory effect induced by these stressors, particularly on mitochondrial proteins and transcripts (Fig. 2). Geneset enrichment analyses revealed upregulation of pathways regulated by activating transcription factors 4/5 (ATF4/5) and the integrated stress response marker CHOP. Analysis of protein expression changes unveiled that 21 proteins were upregulated by both CDDO and MitoBlock-6, including essential mitochondrial chaperones (HSP10 and HSP60). In contrast, the UPRER stressor Tunicamycin did not induce canonical UPRMito activation. Additionally, 142 proteins were commonly downregulated, affecting crucial biological processes such as mitochondrial translation, mitochondrial gene expression, and ribosome biogenesis. Importantly, NAD+ADP-ribosyltransferase activity was highly downregulated, involving PARP proteins with roles in DNA damage repair. On a functional level, mitochondrial stressors led to dose-dependent alterations in HCAEC viability, apoptosis, and reactive oxygen species formation (ROS) measured by resazurin-based assays, caspase 3/7 activity, and MitoSox staining, respectively. Low doses and short incubations with mitochondrial stressors promoted endothelial cell health through cytoprotective UPRMito pathways, while prolonged incubation led to unresolvable mitochondrial stress (exemplarily shown in Fig. 3). SiRNA-mediated gene silencing of the UPRMito regulators ATF4 and ATF5 promoted mitochondrial ROS formation in cells treated with mitochondrial stressors. These effects were particularly more pronounced after ATF5-Knockdown, suggesting that ATF5 is the main regulator of the UPRMito in HCAEC (Fig. 4). Moreover, high-dose UPRMito stressors led to an impairment in mitochondrial membrane integrity with cytosolic release of pro-inflammatory mtDNA. Conclusion UPRMito and UPRER lead to a different stress response. Depending on the underlying conditions, both stress responses can exert both cytoprotective and cytotoxic effects. Further studies are required to analyze the in vivo relevance and possible therapeutic exploitation of the UPRMito in endothelial dysfunction and atherosclerosis.Figures

Enhanced cell stress response and protein degradation capacity underlie artemisinin resistance in Plasmodium falciparum.

Malaria remains a global health burden, killing over half a million people each year. Decreased therapeutic efficacy to artemisinin, the most efficacious antimalarial, has been detected in sub-Saharan Africa, a worrying fact given that over 90% of deaths occur on this continent. Mutations in Kelch13 are the most well-established molecular marker for artemisinin resistance, but these do not explain all artemisinin-resistant isolates. Understanding the biological underpinnings of drug resistance is key to curbing the emergence and spread of artemisinin resistance. Artemisinin-mediated non-specific alkylation leads to the accumulation of misfolded and damaged proteins and activation of the parasite unfolded protein response (UPR). In addition, the parasite proteasome is vital to artemisinin resistance, as we have previously shown that chemical inhibition of the proteasome or mutations in the β2 proteasome subunit increase parasite susceptibility to dihydroartemisinin (DHA), the active metabolite of artemisinins. Here, we investigate parasites with mutations at the Kelch13 and/or 19S and 20S proteasome subunits with regard to UPR regulation and proteasome activity in the context of artemisinin resistance. Our data show that perturbing parasite proteostasis kills parasites, early parasite UPR signaling dictates DHA survival outcomes, and DHA susceptibility correlates with impairment of proteasome-mediated protein degradation. Importantly, we show that functional proteasomes are required for artemisinin resistance in a Kelch13-independent manner, and compound-selective proteasome inhibition demonstrates why artemisinin-resistant Kelch13 mutants remain susceptible to the related antimalarial peroxide OZ439. These data provide further evidence for targeting the parasite proteasome and UPR to overcome existing artemisinin resistance.IMPORTANCEDecreased therapeutic efficacy represents a major barrier to malaria treatment control strategies. The malaria proteasome and accompanying unfolded protein response are crucial to artemisinin resistance, revealing novel antimalarial therapeutic strategies.

The Tumor Suppressor TPD52-Governed Endoplasmic Reticulum Stress is Modulated by APCCdc20.

Aberrant regulation of unfolded protein response (UPR)/endoplasmic reticulum (ER) stress pathway is associated with cancer development, metastasis, and relapse, and the UPR signal transducer ATF6 has been proposed as a diagnostic and prognostic marker for many cancers. However, a causal molecular link between ATF6 activation and carcinogenesis is not established. Here, it is found that tumor protein D52 (TPD52) integrates ER stress and UPR signaling with the chaperone machinery by promoting S2P-mediated cleavage of ATF6. Although TPD52 has been generally considered as an oncogene, TPD52 is identified as a novel tumor suppressor in bladder cancer. Significantly, attenuation of the ER stress via depletion of TPD52 facilitated tumorigenesis in a subset of human carcinomas. Furthermore, the APCCdc20 E3 ligase is validated as the upstream regulator marking TPD52 for polyubiquitination-mediated proteolysis. In addition, inactivation of Cdc20 sensitized cancer cells to treatment with the ER stress inducer in a TPD52-dependent manner. Thus, the study suggests that TPD52 is a novel Cdc20 substrate that may modulate ER stress to prevent tumorigenesis.

CDNF Exerts Anxiolytic, Antidepressant-like, and Procognitive Effects and Modulates Serotonin Turnover and Neuroplasticity-Related Genes.

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.

Structural basis of CDNF interaction with the UPR regulator GRP78

Cerebral dopamine neurotrophic factor (CDNF) is an unconventional neurotrophic factor that is a disease-modifying drug candidate for Parkinson’s disease. CDNF has pleiotropic protective effects on stressed cells, but its mechanism of action remains incompletely understood. Here, we use state-of-the-art advanced structural techniques to resolve the structural basis of CDNF interaction with GRP78, the master regulator of the unfolded protein response (UPR) pathway. Subsequent binding studies confirm the obtained structural model of the complex, eventually revealing the interaction site of CDNF and GRP78. Finally, mutating the key residues of CDNF mediating its interaction with GRP78 not only results in impaired binding of CDNF but also abolishes the neuroprotective activity of CDNF-derived peptides in mesencephalic neuron cultures. These results suggest that the molecular interaction with GRP78 mediates the neuroprotective actions of CDNF and provide a structural basis for development of next generation CDNF-based therapeutic compounds against neurodegenerative diseases.

The unfolded protein response regulates ER exit sites via SNRPB-dependent RNA splicing and contributes to bone development.

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.

Network pharmacology and molecular dynamics study of the effect of the Astragalus-Coptis drug pair on diabetic kidney disease.

Diabetic kidney disease (DKD) is the primary cause of end-stage renal disease. The Astragalus-Coptis drug pair is frequently employed in the management of DKD. However, the precise molecular mechanism underlying its therapeutic effect remains elusive. To investigate the synergistic effects of multiple active ingredients in the Astragalus-Coptis drug pair on DKD through multiple targets and pathways. The ingredients of the Astragalus-Coptis drug pair were collected and screened using the TCMSP database and the SwissADME platform. The targets were predicted using the SwissTargetPrediction database, while the DKD differential gene expression analysis was obtained from the Gene Expression Omnibus database. DKD targets were acquired from the GeneCards, Online Mendelian Inheritance in Man database, and DisGeNET databases, with common targets identified through the Venny platform. The protein-protein interaction network and the "disease-active ingredient-target" network of the common targets were constructed utilizing the STRING database and Cytoscape software, followed by the analysis of the interaction relationships and further screening of key targets and core active ingredients. Gene Ontology (GO) function and Kyoto Ency-clopedia of Genes and Genomes (KEGG) pathway enrichments were performed using the DAVID database. The tissue and organ distributions of key targets were evaluated. PyMOL and AutoDock software validate the molecular docking between the core ingredients and key targets. Finally, molecular dynamics (MD) simulations were conducted to simulate the optimal complex formed by interactions between core ingredients and key target proteins. A total of 27 active ingredients and 512 potential targets of the Astragalus-Coptis drug pair were identified. There were 273 common targets between DKD and the Astragalus-Coptis drug pair. Through protein-protein interaction network topology analysis, we identified 9 core active ingredients and 10 key targets. GO and KEGG pathway enrichment analyses revealed that Astragalus-Coptis drug pair treatment for DKD involves various biological processes, including protein phosphorylation, negative regulation of apoptosis, inflammatory response, and endoplasmic reticulum unfolded protein response. These pathways are mainly associated with the advanced glycation end products (AGE)-receptor for AGE products signaling pathway in diabetic complications, as well as the Lipid and atherosclerosis. Molecular docking and MD simulations demonstrated high affinity and stability between the core active ingredients and key targets. Notably, the quercetin-AKT serine/threonine kinase 1 (AKT1) and quercetin-tumor necrosis factor (TNF) protein complexes exhibited exceptional stability. This study demonstrated that DKD treatment with the Astragalus-Coptis drug pair involves multiple ingredients, targets, and signaling pathways. We propose a novel approach for investigating the molecular mechanism underlying the therapeutic effects of the Astragalus-Coptis drug pair on DKD. Furthermore, we suggest that quercetin is the most potent active ingredient and specifically targets AKT1 and TNF, providing a theoretical foundation for further exploration of pharmacologically active ingredients and elucidating their molecular mechanisms in DKD treatment.

Intranasal Administration of GRP78 Protein (HSPA5) Confers Neuroprotection in a Lactacystin-Induced Rat Model of Parkinson's Disease.

The accumulation of misfolded and aggregated α-synuclein can trigger endoplasmic reticulum (ER) stress and the unfolded protein response (UPR), leading to apoptotic cell death in patients with Parkinson's disease (PD). As the major ER chaperone, glucose-regulated protein 78 (GRP78/BiP/HSPA5) plays a key role in UPR regulation. GRP78 overexpression can modulate the UPR, block apoptosis, and promote the survival of nigral dopamine neurons in a rat model of α-synuclein pathology. Here, we explore the therapeutic potential of intranasal exogenous GRP78 for preventing or slowing PD-like neurodegeneration in a lactacystin-induced rat model. We show that intranasally-administered GRP78 rapidly enters the substantia nigra pars compacta (SNpc) and other afflicted brain regions. It is then internalized by neurons and microglia, preventing the development of the neurodegenerative process in the nigrostriatal system. Lactacystin-induced disturbances, such as the abnormal accumulation of phosphorylated pS129-α-synuclein and activation of the pro-apoptotic GRP78/PERK/eIF2α/CHOP/caspase-3,9 signaling pathway of the UPR, are substantially reversed upon GRP78 administration. Moreover, exogenous GRP78 inhibits both microglia activation and the production of proinflammatory cytokines, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), via the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signaling pathway in model animals. The neuroprotective and anti-inflammatory potential of exogenous GRP78 may inform the development of effective therapeutic agents for PD and other synucleinopathies.

Salicylic acid- and ethylene-dependent effects of the ER stress-inducer tunicamycin on the photosynthetic light reactions in tomato plants

Plant hormones such as ethylene (ET) and salicylic acid (SA) have an elementary role in the regulation of ER stress and unfolded protein response (UPR) in plants via modulating defence responses or inducing oxidative stress. Chloroplasts can be sources and targets of reactive oxygen species (ROS) that affect photosynthetic efficiency, which has not been investigated under tunicamycin (Tm)-induced ER stress. In this study, the direct and indirect effects of Tm on chloroplastic ROS production were first investigated in leaves of wild-type tomato (Solanum lycopersicum L.) plants. Secondly changes in activities of photosystem II and I were analysed under Tm exposure and after application of the chemical chaperone 4-phenylbutyrate (PBA) in different genotypes, focusing on the regulatory role of SA and ET Tm treatments significantly but indirectly induced ROS production in tomato leaves and in parallel it decreased the effective quantum yield of PSII [Y(II)] and PSI [Y(I)], as well as the photochemical quenching coefficient (qP) and the quantum yield of non-photochemical energy dissipation in PSI due to acceptor-side limitation [Y(NA)]. At the same time, Tm increased non-photochemical quenching (NPQ) and cyclic electron flow (CEF) in tomato leaves after 24 h. However, the photosynthetic activity of the SA hydroxylase-overexpressing NahG tomato plants was more severely affected by Tm as compared to wild-type and ET-insensitive Never ripe (Nr) plants. These results suggest the protective role of SA in the regulation of photosynthetic activity contributing to UPR and the survival of plants under ER stress. Interestingly, the activation of photoprotective mechanisms by NPQ was independent of SA but dependent on active ET signalling under ER stress, whereas CEF was reduced by ET due to its higher ratio in Nr plants.

Multi-omics Resources for Understanding Gene Regulation in Response to ER Stress in Plants.

Proteotoxic stress of the endoplasmic reticulum (ER) is a potentially lethal condition that ensues when the biosynthetic capacity of the ER is overwhelmed. A sophisticated and largely conserved signaling, known as the unfolded protein response (UPR), is designed to monitor and alleviate ER stress. In plants, the emerging picture of gene regulation by the UPR now appears to be more complex than ever before, requiring multi-omics-enabled network-level approaches to be untangled. In the past decade, with an increasing access and decreasing costs of next-generation sequencing (NGS) and high-throughput protein-DNA interaction (PDI) screening technologies, multitudes of global molecular measurements, known as omics, have been generated and analyzed by the research community to investigate the complex gene regulation of plant UPR. In this chapter, we present a comprehensive catalog of omics resources at different molecular levels (transcriptomes, protein-DNA interactomes, and networks) along with the introduction of key concepts in experimental and computational tools in data generation and analyses. This chapter will serve as a starting point for both experimentalists and bioinformaticians to explore diverse omics datasets for their biological questions in the plant UPR, with likely applications also in other species for conserved mechanisms.

Identification and functional characterization of activating transcription factor 6 (ATF6) from the mud crab (Scylla paramamosain) in response to hydrogen peroxide and bacterial challenge

Activating transcription factor 6 (ATF6) is critical for regulation of unfolded protein response (UPR), which is involved in the endoplasmic reticulum (ER) proteostasis maintenance and cellular redox regulation. In the present study, a ATF6 gene from the mud crab (designated as Sp-ATF6) has been cloned and identified. The open reading frame of Sp-ATF6 was 1917 bp, encoding a protein of 638 amino acids. The deduced amino acid sequences of Sp-ATF6 contained a typical basic leucine zipper (BZIP domain). Sp-ATF6 was widely expressed in all tested tissues, with the highest expression levels in the hemocytes and the lowest in the muscle. Subcellular localization showed that Sp-ATF6 was expressed in both nucleus and cytoplasm of S2 cells. The expression level of Sp-ATF6 was induced by hydrogen peroxide and V. parahaemolyticus challenge, indicating that the ATF6 pathway was activated in response to ER stress. In order to know more about the regulation mechanism of the Sp-ATF6, RNA interference experiment was investigated. Knocking down Sp-ATF6 in vivo can decrease the expression of antioxidant-related genes (CAT and SOD) and heat shock proteins (HSP90 and HSP70) after V. parahaemolyticus infection. All these results suggested that Sp-ATF6 played a crucial role in the defense against environmental stress and pathogen infection in crustaceans.

GRP78-CAR T cell effector function against solid and brain tumors is controlled by GRP78 expression on T cells

Lack of targetable antigens is a key limitation for developing successful Tcell-based immunotherapies. Members of the unfolded protein response (UPR) represent ideal immunotherapy targets because the UPR regulates the ability of cancer cells to resist cell death, sustain proliferation, and metastasize. Glucose-regulated protein 78 (GRP78) is a key UPR regulator that is overexpressed and translocated to the cell surface of a wide variety of cancers in response to elevated endoplasmic reticulum (ER) stress. We show that GRP78 is highly expressed on the cell surface of multiple solid and brain tumors, making cell surface GRP78 a promising chimeric antigen receptor (CAR) Tcell target. We demonstrate that GRP78-CAR Tcells can recognize and kill GRP78+ brain and solid tumors invitro and invivo. Additionally, our findings demonstrate that GRP78 is upregulated on CAR Tcells upon Tcell activation; however, this expression is tumor-cell-line specific and results in heterogeneous GRP78-CAR Tcell therapeutic response.

Pentapeptide PYRAE triggers ER stress-mediated apoptosis of breast cancer cells in mice by targeting RHBDF1-BiP interaction.

Reinforced cellular responses to endoplasmic reticulum (ER) stress are caused by a variety of pathological conditions including cancers. Human rhomboid family-1 protein (RHBDF1), a multiple transmembrane protein located mainly on the ER, has been shown to promote cancer development, while the binding immunoglobulin protein (BiP) is a key regulator of cellular unfolded protein response (UPR) for the maintenance of ER protein homeostasis. In this study, we investigated the role of RHBDF1 in maintaining ER protein homeostasis in breast cancer cells. We showed that deleting or silencing RHBDF1 in breast cancer cell lines MCF-7 and MDA-MB-231 caused marked aggregation of unfolded proteins in proximity to the ER. We demonstrated that RHBDF1 directly interacted with BiP, and this interaction had a stabilizing effect on the BiP protein. Based on the primary structural motifs of RHBDF1 involved in BiP binding, we found a pentapeptide (PE5) targeted BiP and inhibited BiP ATPase activity. SPR assay revealed a binding affinity of PE5 toward BiP (Kd = 57.7 μM). PE5 (50, 100, 200 μM) dose-dependently promoted ER protein aggregation and ER stress-mediated cell apoptosis in MCF-7 and MDA-MB-231 cells. In mouse 4T1 breast cancer xenograft model, injection of PE5 (10 mg/kg, s.c., every 2 days for 2 weeks) significantly inhibited the tumor growth with markedly increased ER stress and apoptosis-related proteins in tumor tissues. Our results suggest that the ability of RHBDF1 to maintain BiP protein stability is critical to ER protein homeostasis in breast cancer cells, and that the pentapeptide PE5 may serve as a scaffold for the development of a new class of anti-BiP inhibitors.

Aspartyl-tRNA synthetase 2 orchestrates iron-sulfur metabolism in hematopoietic stem cells via fine-tuning alternative RNA splicing

Aspartyl-tRNA synthetase 2 (Dars2) is involved in the regulation of mitochondrial protein synthesis and tissue-specific mitochondrial unfolded protein response (UPRmt). The role of Dars2 in the self-renewal and differentiation of hematopoietic stem cells (HSCs) is unknown. Here, we show that knockout (KO) of Dars2 significantly impairs the maintenance of hematopoietic stem and progenitor cells (HSPCs) without involving its tRNA synthetase activity. Dars2 KO results in significantly reduced expression of Srsf2/3/6 and impairs multiple events of mRNA alternative splicing (AS). Dars2 directly localizes to Srsf3-labeled spliceosomes in HSPCs and regulates the stability of Srsf3. Dars2-deficient HSPCs exhibit aberrant AS of mTOR and Slc22a17. Dars2 KO greatly suppresses the levels of labile ferrous iron and iron-sulfur cluster-containing proteins, which dampens mitochondrial metabolic activity and DNA damage repair pathways in HSPCs. Our study reveals that Dars2 plays a crucial role in the iron-sulfur metabolism and maintenance of HSPCs by modulating RNA splicing.

Unfolded Protein Response Signaling in Liver Disorders: A 2023 Updated Review.

Endoplasmic reticulum (ER) is the site for synthesis and folding of secreted and transmembrane proteins. Disturbance in the functioning of ER leads to the accumulation of unfolded and misfolded proteins, which finally activate the unfolded protein response (UPR) signaling. The three branches of UPR-IRE1 (Inositol requiring enzyme 1), PERK (Protein kinase RNA-activated (PKR)-like ER kinase), and ATF6 (Activating transcription factor 6)-modulate the gene expression pattern through increased expression of chaperones and restore ER homeostasis by enhancing ER protein folding capacity. The liver is a central organ which performs a variety of functions which help in maintaining the overall well-being of our body. The liver plays many roles in cellular physiology, blood homeostasis, and detoxification, and is the main site at which protein synthesis occurs. Disturbance in ER homeostasis is triggered by calcium level imbalance, change in redox status, viral infection, and so on. ER dysfunction and subsequent UPR signaling participate in various hepatic disorders like metabolic (dysfunction) associated fatty liver disease, liver cancer, viral hepatitis, and cholestasis. The exact role of ER stress and UPR signaling in various liver diseases is not fully understood and needs further investigation. Targeting UPR signaling with drugs is the subject of intensive research for therapeutic use in liver diseases. The present review summarizes the role of UPR signaling in liver disorders and describes why UPR regulators are promising therapeutic targets.

CD44 correlates with longevity and enhances basal ATF6 activity and ER stress resistance

The naked mole rat (NMR) is the longest-lived rodent, resistant to multiple age-related diseases including neurodegeneration. However, the mechanisms underlying the NMR's resistance to neurodegenerative diseases remain elusive. Here, we isolated oligodendrocyte progenitor cells (OPCs) from NMRs and compared their transcriptome with that of other mammals. Extracellular matrix (ECM) genes best distinguish OPCs of long- and short-lived species. Notably, expression levels of CD44, an ECM-binding protein that has been suggested to contribute to NMR longevity by mediating the effect of hyaluronan (HA), are not only high in OPCs of long-lived species but also positively correlate with longevity in multiple cell types/tissues. We found that CD44 localizes to the endoplasmic reticulum (ER) and enhances basal ATF6 activity. CD44 modifies proteome and membrane properties of the ER and enhances ER stress resistance in a manner dependent on unfolded protein response regulators without the requirement of HA. HA-independent role of CD44 in proteostasis regulation may contribute to mammalian longevity.

An IRE1-proteasome system signalling cohort controls cell fate determination in unresolved proteotoxic stress of the plant endoplasmic reticulum.

Excessive accumulation of misfolded proteins in the endoplasmic reticulum (ER) causes ER stress, which is an underlying cause of major crop losses and devastating human conditions. ER proteostasis surveillance is mediated by the conserved master regulator of the unfolded protein response (UPR), Inositol Requiring Enzyme 1 (IRE1), which determines cell fate by controlling pro-life and pro-death outcomes through as yet largely unknown mechanisms. Here we report that Arabidopsis IRE1 determines cell fate in ER stress by balancing the ubiquitin-proteasome system (UPS) and UPR through the plant-unique E3 ligase, PHOSPHATASE TYPE 2CA (PP2CA)-INTERACTING RING FINGER PROTEIN 1 (PIR1). Indeed, PIR1 loss leads to suppression of pro-death UPS and the lethal phenotype of an IRE1 loss-of-function mutant in unresolved ER stress in addition to activating pro-survival UPR. Specifically, in ER stress, PIR1 loss stabilizes ABI5, a basic leucine zipper (bZIP) transcription factor, that directly activates expression of the critical UPR regulator gene, bZIP60, triggering transcriptional cascades enhancing pro-survival UPR. Collectively, our results identify new cell fate effectors in plant ER stress by showing that IRE1's coordination of cell death and survival hinges on PIR1, a key pro-death component of the UPS, which controls ABI5, a pro-survival transcriptional activator of bZIP60.

Differentially Expressed Genes Regulating Glutathione Metabolism, Protein-Folding, and Unfolded Protein Response in Pancreatic β-Cells in Type 2 Diabetes Mellitus.

Impaired redox homeostasis in the endoplasmic reticulum (ER) may contribute to proinsulin misfolding and thus to activate the unfolded protein response (UPR) and apoptotic pathways, culminating in pancreatic β-cell loss and type 2 diabetes (T2D). The present study was designed to identify differentially expressed genes (DEGs) encoding enzymes for glutathione metabolism and their impact on the expression levels of genes regulating protein folding and UPR in β-cells of T2D patients. The GEO transcriptome datasets of β-cells of diabetics and non-diabetics, GSE20966 and GSE81608, were analyzed for 142 genes of interest using limma and GREIN software, respectively. Diabetic β-cells showed dataset-specific patterns of DEGs (FDR ≤ 0.05) implicated in the regulation of glutathione metabolism (ANPEP, PGD, IDH2, and CTH), protein-folding (HSP90AB1, HSP90AA1, HSPA1B, HSPA8, BAG3, NDC1, NUP160, RLN1, and RPS19BP1), and unfolded protein response (CREB3L4, ERP27, and BID). The GCLC gene, encoding the catalytic subunit of glutamate-cysteine ligase, the first rate-limiting enzyme of glutathione biosynthesis, was moderately down-regulated in diabetic β-cells from both datasets (p ≤ 0.05). Regression analysis established that genes involved in the de novo synthesis of glutathione, GCLC, GCLM, and GSS affect the expression levels of genes encoding molecular chaperones and those involved in the UPR pathway. This study showed for the first time that diabetic β-cells exhibit alterations in the expression of genes regulating glutathione metabolism, protein-folding, and UPR and provided evidence for the molecular crosstalk between impaired redox homeostasis and abnormal protein folding, underlying ER stress in type 2 diabetes.

Overexpression of VIRMA confers vulnerability to breast cancers via the m6A-dependent regulation of unfolded protein response

Virilizer-like m6A methyltransferase-associated protein (VIRMA) maintains the stability of the m6A writer complex. Although VIRMA is critical for RNA m6A deposition, the impact of aberrant VIRMA expression in human diseases remains unclear. We show that VIRMA is amplified and overexpressed in 15–20% of breast cancers. Of the two known VIRMA isoforms, the nuclear-enriched full-length but not the cytoplasmic-localised N-terminal VIRMA promotes m6A-dependent breast tumourigenesis in vitro and in vivo. Mechanistically, we reveal that VIRMA overexpression upregulates the m6A-modified long non-coding RNA, NEAT1, which contributes to breast cancer cell growth. We also show that VIRMA overexpression enriches m6A on transcripts that regulate the unfolded protein response (UPR) pathway but does not promote their translation to activate the UPR under optimal growth conditions. Under stressful conditions that are often present in tumour microenvironments, VIRMA-overexpressing cells display enhanced UPR and increased susceptibility to death. Our study identifies oncogenic VIRMA overexpression as a vulnerability that may be exploited for cancer therapy.

Omics analyses in citrus reveal a possible role of RNA translation pathways and Unfolded Protein Response regulators in the tolerance to combined drought, high irradiance, and heat stress.

Environmental changes derived from global warming and human activities increase the intensity and frequency of stressful conditions for plants. Multiple abiotic factors acting simultaneously enhance stress pressure and drastically reduce plant growth, yield, and survival. Stress combination causes a specific stress situation that induces a particular plant response different to the sum of responses to the individual stresses. Here, by comparing transcriptomic and proteomic profiles to different abiotic stress combinations in two citrus genotypes, Carrizo citrange (Citrus sinensis×Poncirus trifoliata) and Cleopatra mandarin (Citrus reshni), with contrasting tolerance to different abiotic stresses, we revealed key responses to the triple combination of heat stress, high irradiance and drought. The specific transcriptomic response to this stress combination in Carrizo was directed to regulate RNA metabolic pathways and translation processes, potentially conferring an advantage with respect to Cleopatra. In addition, we found endoplasmic reticulum stress response as common to all individual and combined stress conditions in both genotypes and identified the accumulation of specific groups of heat shock proteins (HSPs), such as small HSPs and HSP70s, and regulators of the unfolded protein response, BiP2 and PDIL2-2, as possible factors involved in citrus tolerance to triple stress combination. Taken together, our findings provide new insights into the acclimation process of citrus plants to multiple stress combination, necessary for increasing crop tolerance to the changing climatic conditions.