Hexavalent chromium Cr(VI) exists widely in occupational environments. The mechanistic target of rapamycin (mTOR) has been well-documented to regulate autophagy negatively. However, we found that low ...concentration of Cr(VI) (0.2 μM) elevated both mTOR and autophagy and promote cell survival. Conversely, high concentration of Cr(VI) (6 μM) caused cell death by inhibiting mTOR and subsequently inducing autophagy. Tunicamycin (Tm), as an Endoplasmic reticulum (ER) stress activator was used to induce mild ER stress at 0.1 μg/ml and it activated both autophagy and mTOR, which also caused cell migration in a similar manner to that observed with low concentration of Cr(VI). Severe ER stress caused by Tm (2 μg/ml) decreased mTOR, increased autophagy and then inhibited cell migration, which was the same as 6 μM Cr(VI) treatment, although Cr(VI) in high concentration inhibited ER stress. Activating transcription factor 4 (ATF4), a downstream target of ER stress, only increased under mild ER stress but decreased under severe ER stress and 6 μM Cr(VI) treatment. Chromatin immunoprecipitation (ChIP) experiment indicated that ATF4 could bind to the promoter of ATG4B and AKT1. To sum up, our data revealed that mild ER stress induced by low concentration of Cr(VI) could enhance transcriptional regulation of ATG4B and AKT1 by ATF4, which induced both autophagy and mTOR to promote cell viability.
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•Cr(VI) simultaneously induced autophagy and mTOR in vitro and in vivo.•High concentration of Tm and Cr(VI) inhibited mTOR, induced autophagy but reduced cell migration.•Low concentration of Cr(VI)-induced mild ER stress accelerated cell survival by increasing ATF4-mediated autophagy and mTOR.•ATF4 regulated autophagy by binding to ATG4B promoter and ATF4 increased mTOR by binding to AKT1 promoter.
An accumulation of unfolded or misfolded proteins in the endoplasmic reticulum (ER) leads to stress conditions. To mitigate such circumstances, stressed cells activate a homeostatic intracellular ...signaling network cumulatively called the unfolded protein response (UPR), which orchestrates the recuperation of ER function. Macroautophagy (hereafter autophagy), an intracellular lysosome-mediated bulk degradation pathway for recycling and eliminating wornout proteins, protein aggregates, and damaged organelles, has also emerged as an essential protective mechanism during ER stress. These 2 systems are dynamically interconnected, and recent investigations have revealed that ER stress can either stimulate or inhibit autophagy. However, the stress-associated molecular cues that control the changeover switch between induction and inhibition of autophagy are largely obscure. This review summarizes the crosstalk between ER stress and autophagy and their signaling networks mainly in mammalian-based systems. Additionally, we highlight current knowledge on selective autophagy and its connection to ER stress.
Unfolded Protein Response in Arabidopsis Ruberti, Cristina; Brandizzi, Federica
Methods in molecular biology (Clifton, N.J.),
2024, Volume:
2772
Journal Article
Open access
The unfolded protein response (UPR) is a highly regulated signaling pathway that is largely conserved across eukaryotes. It is essential for cell homeostasis under environmental and physiological ...conditions that perturb the protein folding in the endoplasmic reticulum (ER). Arabidopsis is one of the outstanding multicellular model systems in which to investigate the UPR. Here, we described a protocol to induce the UPR in plants, specifically Arabidopsis, and to estimate their ability to cope with ER stress through the quantification of physiological parameters.
The role of ER in the proper synthesis and correct folding of protein, to establish the homeostasis of organism, is well documented by various studies. Following excessive oxidative stress, the ...accumulation of unfolded/misfolded proteins increase in the lumen of ER which blocks the inhibitory effect of GRP78 on UPR regulatory proteins; IRE1, PERK and ATF6.
It is clear that ER stress mediated UPR is a potent inducer of proteasomal and autophagic activities which serves as a degradation process for maintaining stress tolerance, limited damage and viability under stress conditions. Interestingly, aberrantly activation of autophagy can result in destroying major portions of the cytosol and organelles, which will lead to autophagic death of cardiac myocytes, and development of cardiac failure.
In the present study Grp78, Grp94, pIRE1, pPERK, derlin-1, proteasome β5 subunit, VCP, beclin-1 and LC3-II, well known markers of ER stress, proteasomal and autophagic activities, are investigated in the heart tissue of rabbits. Furthermore, autophagic activity in heart tissue is observed in hypercholesterolemic vs control rabbits by electron microscopy. Additionally, colocalization studies of mitochondria and p62 in cryo sections of heart tissue were applied to understand the levels of mitochondria degradation by autophagy.
Uvaol (UV), a pentacyclic triterpene found in olives and virgin olive oil, is known for its anti-inflammatory and antioxidant effects in various disease models. While olive oil is reported to reduce ...obesity and insulin resistance, the specific impact of UV on liver lipid metabolism and its molecular mechanisms are not fully understood. In this study, hepatic lipid accumulation was measured using oil red O staining, and protein expression levels in liver cells were assessed via Western blot analysis. Apoptosis was evaluated through cell viability and caspase 3 activity assays. UV treatment reduced lipid accumulation, fatty acid uptake, apoptosis, and ER stress in palmitate-treated liver cells. Additionally, UV enhanced fatty acid oxidation. Mechanistically, increased SIRT6 expression and autophagy were observed in UV-treated cells. SIRT6-targeted siRNA or 3-methyladenine blocked the effects of UV in hyperlipidemic cells. In conclusion, UV improves SIRT6/autophagy signaling, reducing lipid deposition and apoptosis in liver cells under high lipid conditions. This in vitro study provides strong evidence for potential therapeutic strategies for hepatic steatosis.
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•Uvaol (UV) attenuates palmitate-induced lipogenic lipid deposition and apoptosis in hepatocytes.•UV suppresses PTP1B expression and ER stress in palmitate-treated hepatocytes.•UV enhances SIRT6/autophagy pathway.•"SIRT6/autophagy pathway reduces UV-induced lipogenic lipid accumulation in hepatocytes by inhibiting PTP1B/ER stress."
•CFRD is characterised by exocrine pancreatic, islet loss, and insulin insufficiency.•Mechanistic gaps and conflicting data present in current understanding of CFRD.•Oxidative and endoplasmic ...reticulum stress may play a key role in CF pancreatic damage.•Exosomal microRNA could amplify exocrine-endocrine cross-talk and damage in CFRD.
CF-related diabetes (CFRD) is a prevalent comorbidity in people with Cystic Fibrosis (CF), significantly impacting morbidity and mortality rates. This review article critically evaluates the current understanding of CFRD molecular mechanisms, including the role of CFTR protein, oxidative stress, unfolded protein response (UPR) and intracellular communication. CFRD manifests from a complex interplay between exocrine pancreatic damage and intrinsic endocrine dysfunction, further complicated by the deleterious effects of misfolded CFTR protein on insulin secretion and action. Studies indicate that ER stress and subsequent UPR activation play critical roles in both exocrine and endocrine pancreatic cell dysfunction, contributing to β-cell loss and insulin insufficiency. Additionally, oxidative stress and altered calcium flux, exacerbated by CFTR dysfunction, impair β-cell survival and function, highlighting the significance of antioxidant pathways in CFRD pathogenesis.
Emerging evidence underscores the importance of exosomal microRNAs (miRNAs) in mediating inflammatory and stress responses, offering novel insights into CFRD's molecular landscape. Despite insulin therapy remaining the cornerstone of CFRD management, the variability in response to CFTR modulators underscores the need for personalized treatment approaches. The review advocates for further research into non-CFTR therapeutic targets, emphasizing the need to address the multifaceted pathophysiology of CFRD. Understanding the intricate mechanisms underlying CFRD will pave the way for innovative treatments, moving beyond insulin therapy to target the disease's root causes and improve the quality of life for individuals with CF.
Disturbances in endoplasmic reticulum (ER) function lead to ER stress which, when severe or prolonged, may result in apoptosis. Severe ER stress has been implicated in several pathological conditions ...including pre-eclampsia, a multisystem disorder of pregnancy associated with the release of pro-inflammatory factors from the placenta into the maternal circulation. Here, we show that severe ER stress induced by two distinct mechanisms in BeWo choriocarcinoma cells leads to the release of extracellular vesicles (EVs) carrying pro-inflammatory damage-associated molecular pattern (DAMP) molecules. Co-treatment with the antioxidant pyrrolidine dithiocarbamate results in a reduction in ER stress-induced EV-associated DAMP release. We further demonstrate that severe ER stress is associated with changes in the expression of several stress-related proteins, notably Cited-2 and phosphorylated JNK. Together, these data indicate that severe ER stress-mediated release of EV-associated DAMPs may contribute to the heightened systemic maternal inflammatory response characteristic of pre-eclampsia and may also be relevant to other chronic inflammatory diseases which display elevated ER stress.
Periprosthetic osteolysis and subsequent aseptic loosening are the primary causes of failure following total joint arthroplasty. Wear particle–induced osteogenic impairment is recognized as an ...important contributing factor in the development of osteolysis, with endoplasmic reticulum (ER) stress emerging as a pivotal underlying mechanism. Hence, searching for potential therapeutic targets and agents capable of modulating ER stress in osteoblasts is crucial for preventing aseptic loosening. Kaempferol (KAE), a natural flavonol compound, has shown promising osteoprotective effects and anti-ER stress properties in diverse diseases. However, the influence of KAE on ER stress-mediated osteogenic impairment induced by wear particles remains unclear. In this study, we observed that KAE effectively relieved TiAl6V4 particles–induced osteolysis by improving osteogenesis in a mouse calvarial model. Furthermore, we demonstrated that KAE could attenuate ER stress-mediated apoptosis in osteoblasts exposed to TiAl6V4 particles, both in vitro and in vivo. Mechanistically, our results revealed that KAE mitigated ER stress-mediated apoptosis by upregulating the IRE1α–XBP1s pathway while concurrently partially inhibiting the IRE1α-regulated RIDD and JNK activation. Collectively, our findings suggest that KAE is a prospective therapeutic agent for treating wear particle–induced osteolysis and highlight the IRE1α–XBP1s pathway as a potential therapeutic target for preventing aseptic loosening.
Copper is an essential trace element, and excessive exposure could result in hepatoxicity, however, the underlying molecular mechanisms remain incompletely understood. The present study is aimed to ...investigate the molecular mechanisms of copper sulfate (CuSO4) exposure-induced hepatoxicity both in vivo and in vitro. In vitro, HepG2 and L02 cells were exposed to various doses of CuSO4 for 24 h. Cell viability, ROS production, oxidative stress biomarkers, mitochondrial functions, ultrastructure, intracellular calcium (Ca2+) concentration, and the expression of proteins related to mitochondrial apoptosis and endoplasmic reticulum (ER) stress were assessed. In vivo, C57BL/6 mice were treated with CuSO4 at doses of 10 and 30 mg/kg BW/day and co-treated with 4-PBA at 100 mg/kg BW/day for 35 days. Subsequently, liver function, histopathological features, and protein expression were evaluated. Results found that exposure to CuSO4 at concentrations of 100–400 μM for 24 h significantly decreased the viabilities of HepG2 and L02 cells and it was in a dose-dependent manner. Additionally, CuSO4 exposure induced significant oxidative stress and mitochondrial dysfunction in HepG2 cells, which were partially ameliorated by the antioxidant N-acetylcysteine (NAC). Furthermore, CuSO4 exposure prominently triggered ER stress, as evidenced by the upregulation of GRP94, GRP78, phosphorylated forms of PERK and eIF2α, and CHOP proteins in livers of mice and HepG2 cells. NAC treatment significantly inhibited CuSO4 exposure -induced ER stress in HepG2 cells. Pharmacological inhibition of ER stress through co-treatment with 4-PBA and the PERK inhibitor GSK2606414, as well as genetic knockdown of ATF4, partially mitigated CuSO4-induced cytotoxicity in HepG2 cells by reducing mitochondrial dysfunction and inhibiting the mitochondrial apoptotic pathway. Moreover, 4-PBA treatment significantly attenuated CuSO4-induced caspase activation and hepatoxicity in mice. In conclusion, these results reveal that CuSO4-induced hepatotoxicity involves mitochondrial dysfunction and ER stress by activating oxidative stress induction and PERK/ATF4 pathway.
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•Copper exposure could activate the PERK/ATF4/CHOP pathway to promote ER stress.•Copper exposure causes oxidative stress and mitochondrial dysfunction in vivo and in vitro.•Copper exposure-caused mitochondrial damage involves ROS production and ER stress.•Copper exposure could trigger the loss of mitochondrial CI and CII via ATF4.•Targeting ER stress could attenuate Cu exposure-induced cytotoxicity and hepatoxicity.
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