Fragmentation Of Endoplasmic Reticulum Research Articles

Single fluorescent probe reveals glutathione-viscosity crosstalk in the endoplasmic reticulum during ferroptosis and extracellular vesicles in synergistic ferroptosis-apoptosis

Ferroptosis is an iron-dependent cell death triggered by lipid peroxidation accumulation, representing a redox imbalance in cells. The increased knowledge of ferroptosis has led to various potential strategies to develop new cancer therapies, such as synergistic ferroptosis-apoptosis. The endoplasmic reticulum (ER) is the major hub for lipid metabolism in cells and plays a central role in lipid peroxidation. Glutathione (GSH) and viscosity levels in ER are important parameters for intracellular redox homeostasis, which remain unclear in ferroptosis and synergistic ferroptosis-apoptosis. This study developed an ER-targeting fluorescent probe (MMPY) with a dual-emission response to GSH and viscosity. Fluorescent cell imaging using MMPY revealed different GSH-viscosity crosstalk in the ER for the four ferroptosis pathways induced by erastin, RSL3, FIN56, and FINO2. GSH and viscosity changes in the ER during synergistic ferroptosis-apoptosis were also investigated. Interestingly, the new types of extracellular vesicles were observed during the synergistic process, which burdened the ER fragments with different GSH contents.

Metallic and metallic oxide nanoparticles toxicity primarily targets the mitochondria of hepatocytes and renal cells.

Nanoparticles (NPs) are utilized in various applications, posing potential risks to human health, tissues, cells, and macromolecules. This study aimed to investigate the ultrastructural alterations in hepatocytes and renal tubular cells induced by metallic and metal oxide NPs. Adult healthy male Wistar albino rats (Rattus norvegicus) were divided into 6 (n = 7) control and 6 treated groups (n = 7). The rats in the treated groups exposed daily to silver NPs, gold NPs, zinc oxide NPs, silicon dioxide NPs, copper oxide NPs, and ferric oxide NPs for 35 days. The members of the control group for each corresponding NPs received the respective vehicle. Liver and kidney tissue blocks from all rats were processed for Transmission Electron Microscopy (TEM) examinations. The hepatocytes and renal tubular cells of all NPs-treated rats demonstrated mitochondrial ultrastructural alterations mainly cristolysis, swelling, membrane disruption, lucent matrices, matrices lysis, and electron-dense deposits. However, other organelles demonstrated injury but to a lesser extent in the form of shrunken nuclei, nuclear membrane indentation, endoplasmic reticulum fragmentation, cellular membranes enfolding, brush border microvilli disruption, lysosomal hyperplasia, ribosomes dropping, and peroxisome formation. One may conclude from the findings that the hepatocytes and the renal tubular cells mitochondria are the main targets for nanoparticles toxicity ending in mitochondrial disruption and cell injury. Further studies taking into account the relation of mitochondrial ultrastructural damage with a weakened antioxidant defense system induced by chronic exposure to nanomaterials are needed.

ER-phagy restrains inflammatory responses through its receptor UBAC2

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.

Massive ER protein disposal by reticulophagy receptors and selective disposal by TOLLIP

ABSTRACT Proteostasis of the endoplasmic reticulum (ER) is maintained by coordinated action of two major catabolic pathways: proteasome-dependent ER-associated degradation (ERAD) and less characterized lysosomal pathways. Recent studies on ER-specific autophagy (termed “reticulophagy”) have highlighted the importance of lysosomes for ER proteostasis. Key to this process are proteins termed reticulophagy receptors that connect ER fragments and Atg8-family proteins, facilitating the lysosomal degradation of both native and aberrant ER proteins in a relatively nonselective manner. In contrast, our recent work identified TOLLIP as a novel type of cargo receptor specifically dedicated to the lysosomal degradation of aberrant ER membrane proteins. The clients of TOLLIP include an engineered model substrate, which mimics an ER-retained aberrant membrane protein, and motor neuron disease-linked misfolded mutants of VAPB and BSCL2/Seipin. TOLLIP acts as a receptor to connect these aberrant ER membrane proteins and phosphatidylinositol-3-phosphate (PtdIns3P) by recognizing the former through its misfolding-sensing intrinsically disordered region (IDR) and ubiquitin-binding CUE domain, and the latter through its C2 domain. These interactions enable PtdIns3P-dependent vesicular trafficking of aberrant membrane proteins to lysosomes without promoting reticulophagic turnover of bulk ER.

Rab18 Drift in Lipid Droplet and Endoplasmic Reticulum Interactions of Adipocytes under Obesogenic Conditions.

The adipose tissue stores excess energy in the form of neutral lipids within adipocyte lipid droplets (LDs). The correct function of LDs requires the interaction with other organelles, such as the endoplasmic reticulum (ER) as well as with LD coat-associated proteins, including Rab18, a mediator of intracellular lipid trafficking and ER-LD interaction. Although perturbations of the inter-organelle contact sites have been linked to several diseases, such as cancer, no information regarding ER-LD contact sites in dysfunctional adipocytes from the obese adipose tissue has been published to date. Herein, the ER-LD connection and Rab18 distribution at ER-LD contact sites are examined in adipocytes challenged with fibrosis and inflammatory conditions, which represent known hallmarks of the adipose tissue in obesity. Our results show that adipocytes differentiated in fibrotic conditions caused ER fragmentation, the expansion of ER-LD contact sites, and modified Rab18 dynamics. Likewise, adipocytes exposed to inflammatory conditions favored ER-LD contact, Rab18 accumulation in the ER, and Rab18 redistribution to large LDs. Finally, our studies in human adipocytes supported the suggestion that Rab18 transitions to the LD coat from the ER. Taken together, our results suggest that obesity-related pathogenic processes alter the maintenance of ER-LD interactions and interfere with Rab18 trafficking through these contact sites.

8-OR: Novel Mechanism of Lipoprotein Lipase Folding and Maturation in Adipocytes

Lipoprotein lipase (LPL) plays a central role in systemic lipid clearance by catalyzing intravascular hydrolysis of lipoprotein triglycerides, hence representing an important target in the treatment of hyperlipidemia in diabetes. Nascent LPL protein undergoes folding and maturation in the endoplasmic reticulum (ER), with only the properly folded ones exiting the ER and reaching to the cell surface. However, the maturation process for nascent LPL in the ER remains largely unknown. Here we show that LPL maturation in the ER is controlled by compensatory actions of two ER protein quality control mechanisms, ER-associated degradation (ERAD) and ER-phagy, in adipocytes. Specifically, we show that LPL is misfolding prone and degraded by ERAD, and when ERAD is compromised, LPL forms aggregates which is cleared by ER-phagy. In adipocytes lacking both ERAD and ER-phagy, ER fragments containing LPL aggregates cluster and fuse to form a novel cellular architecture, termed as Coalescence of ER Fragments (CERFs), where LPL undergo soluble-to-insoluble phase transition catalyzed by disulfide bond-mediated interactions. Lastly, mice with adipocyte specific ERAD and ER-phagy deficiency exhibited postprandial hyperlipidemia. In summary, this study provides novel insights into not only LPL biogenesis, but also the crosstalk between protein quality control pathways in the maintenance of ER homeostasis, as well as the disease pathogenesis, more broadly, associated with protein folding defects in the ER. Disclosure S.Wu: None. L.Qi: None.

The mechanisms to dispose of misfolded proteins in the endoplasmic reticulum of adipocytes

Endoplasmic reticulum (ER)-associated degradation (ERAD) and ER-phagy are two principal degradative mechanisms for ER proteins and aggregates, respectively; however, the crosstalk between these two pathways under physiological settings remains unexplored. Using adipocytes as a model system, here we report that SEL1L-HRD1 protein complex of ERAD degrades misfolded ER proteins and limits ER-phagy and that, only when SEL1L-HRD1 ERAD is impaired, the ER becomes fragmented and cleared by ER-phagy. When both are compromised, ER fragments containing misfolded proteins spatially coalesce into a distinct architecture termed Coalescence of ER Fragments (CERFs), consisted of lipoprotein lipase (LPL, a key lipolytic enzyme and an endogenous SEL1L-HRD1 substrate) and certain ER chaperones. CERFs enlarge and become increasingly insoluble with age. Finally, we reconstitute the CERFs through LPL and BiP phase separation in vitro, a process influenced by both redox environment and C-terminal tryptophan loop of LPL. Hence, our findings demonstrate a sequence of events centered around SEL1L-HRD1 ERAD to dispose of misfolded proteins in the ER of adipocytes, highlighting the profound cellular adaptability to misfolded proteins in the ER in vivo.

Role of endoplasmic reticulum autophagy in acute lung injury.

Acute lung injury (ALI) and acute respiratory distress syndrome (ARDS), the prime causes of morbidity and mortality in critically ill patients, are usually treated by general supportive treatments. Endoplasmic reticulum autophagy (ER-phagy) maintains cellular homeostasis by degrading damaged endoplasmic reticulum (ER) fragments and misfolded proteins. ER-phagy is crucial for maintaining ER homeostasis and improving the internal environment. ER-phagy has a particular role in some aspects, such as immunity, inflammation, cell death, pathogen infection, and collagen quality. In this review, we summarized the definition, epidemiology, and pathophysiology of ALI/ARDS and described the regulatory mechanisms and functions of ER-phagy as well as discussed the potential role of ER-phagy in ALI/ARDS from the perspectives of immunity, inflammation, apoptosis, pathogen infection, and fibrosis to provide a novel and effective target for improving the prognosis of ALI/ARDS.

The role of selective autophagy in pathogen infection

<p indent="0mm">Autophagy is a highly conserved physiological process of material degradation and recycling. According to the different pathways of degrading cargo, autophagy is classified into three types: Macroautophagy, microautophagy and chaperone-mediated autophagy (CMA). Macroautophagy refers to the process in which cargos to be degraded are wrapped in double-membrane autophagosome and transported to lysosomes for degradation. Microautophagy refers to the process in which lysosomal membrane invagination or protrusion wraps the cargo to be degraded and directly degrades it. Chaperone-mediated autophagy refers to the process in which molecular chaperones restore intracellular proteins from the folded state to the unfolded state, and then transport them to lysosomes for degradation. Autophagy was previously defined as a non-selective process induced under starvation conditions. However, developments in autophagy-related research and in-depth studies have revealed that autophagy can be either a non-selective or selective process. Selective autophagy refers to the selective cellular degradation process that targets and degrades dysfunctional organelles, misfolded proteins, or invading pathogens. According to the different targets of degradation, selective autophagy is divided into mitophagy, endoplasmic reticulum autophagy (ER-phagy), proteasome autophagy (proteaphagy), ribosomal autophagy (ribophagy), lipophagy, nucleophagy, and xenophagy, and so on. Currently, mitophagy, ER-phagy, and xenophagy are hot research topics. Mitophagy is an autophagic process that selectively targets and degrades damaged or redundant mitochondria and is essential for mitochondrial quality control and maintenance of cell homeostasis. In recent years, accumulating studies have shown that some viruses or bacteria regulate the mitophagy process by various direct or indirect strategies, thereby weakening innate immune responses such as type I interferon response, apoptosis, and the activation of inflammasome to facilitate infection. Further, ER-phagy is a selective autophagic process that targets and degrades damaged endoplasmic reticulum fragments and plays essential roles in endoplasmic reticulum quality control and maintenance of cellular homeostasis. The on-going studies on ER-phagy mechanism are limited compared with those on mitophagy mechanism, but studies have reported that ER-phagy plays a vital role in the immune escape of pathogens. Xenophagy refers to the process in which eukaryotic cells recognize and degrade invading pathogens through selective autophagy. However, some studies have reported that some invading pathogens such as <italic>Mycobacterium tuberculosis</italic> and <italic>Salmonella</italic> inhibit xenophagy in host to achieve intracellular survival. Selective autophagy is a process that targets different cargo that is to be degraded and shares some crucial autophagy-related proteins. When multiple selective autophagy processes simultaneously occur, these processes compete for the shared proteins. However, studies on the crosstalk between different selective autophagy processes are limited. In summary, this article reviews the molecular mechanisms of mitophagy, ER-phagy, and xenophagy and their roles in pathogen infection to provide a reference for selective autophagy-related studies.

Phosphorylation of FAM134C by CK2 controls starvation-induced ER-phagy.

Selective degradation of the endoplasmic reticulum (ER) via autophagy (ER-phagy) is initiated by ER-phagy receptors, which facilitate the incorporation of ER fragments into autophagosomes. FAM134 reticulon family proteins (FAM134A, FAM134B, and FAM134C) are ER-phagy receptors with structural similarities and nonredundant functions. Whether they respond differentially to the stimulation of ER-phagy is unknown. Here, we describe an activation mechanism unique to FAM134C during starvation. In fed conditions, FAM134C is phosphorylated by casein kinase 2 (CK2) at critical residues flanking the LIR domain. Phosphorylation of these residues negatively affects binding affinity to the autophagy proteins LC3. During starvation, mTORC1 inhibition limits FAM134C phosphorylation by CK2, hence promoting receptor activation and ER-phagy. Using a novel tool to study ER-phagy in vivo and FAM134C knockout mice, we demonstrated the physiological relevance of FAM134C phosphorylation during starvation-induced ER-phagy in liver lipid metabolism. These data provide a mechanistic insight into ER-phagy regulation and an example of autophagy selectivity during starvation.

Nicotine in Combination with SARS-CoV-2 Affects Cells Viability, Inflammatory Response and Ultrastructural Integrity.

The aims of our study are to: (i) investigate the ability of nicotine to modulate the expression level of inflammatory cytokines in A549 cells infected with SARS-CoV-2; (ii) elucidate the ultrastructural features caused by the combination nicotine+SARS-CoV-2; and (iii) demonstrate the mechanism of action. In this study, A549 cells pretreated with nicotine were either exposed to LPS or poly(I:C), or infected with SARS-CoV-2. Treated and untreated cells were analyzed for cytokine production, cytotoxicity, and ultrastructural modifications. Vero E6 cells were used as a positive reference. Cells pretreated with nicotine showed a decrease of IL6 and TNFα in A549 cells induced by LPS or poly(I:C). In contrast, cells exposed to SARS-CoV-2 showed a high increase of IL6, IL8, IL10 and TNFα, high cytopathic effects that were dose- and time-dependent, and profound ultrastructural modifications. These modifications were characterized by membrane ruptures and fragmentation, the swelling of cytosol and mitochondria, the release of cytoplasmic content in extracellular spaces (including osmiophilic granules), the fragmentation of endoplasmic reticulum, and chromatin disorganization. Nicotine increased SARS-CoV-2 cytopathic effects, elevating the levels of inflammatory cytokines, and inducing severe cellular damage, with features resembling pyroptosis and necroptosis. The protective role of nicotine in COVID-19 is definitively ruled out.

ER-Phagy: Quality and Quantity Control of the Endoplasmic Reticulum by Autophagy.

The endoplasmic reticulum (ER) is the largest organelle and has multiple roles in various cellular processes such as protein secretion, lipid synthesis, calcium storage, and organelle biogenesis. The quantity and quality of this organelle are controlled by the ubiquitin-proteasome system and autophagy (termed "ER-phagy"). ER-phagy is defined as the degradation of part of the ER by the vacuole or lysosomes, and there are at least two types of ER-phagy: macro-ER-phagy and micro-ER-phagy. In macro-ER-phagy, ER fragments are enclosed by autophagosomes, which is mediated by ER-phagy receptors. In micro-ER-phagy, a portion of the ER is engulfed directly by the vacuole or lysosomes. In these two pathways, some proteins in the ER lumen can be recognized selectively and subjected to ER-phagy. This review summarizes our current knowledge of ER-phagy, focusing on its membrane dynamics, molecular mechanisms, substrate specificity, and physiological significance.

Structural alterations in cell organelles induced by photodynamic treatment with chlorin p6 -histamine conjugate in human oral carcinoma cells probed by 3D fluorescence microscopy.

We have explored the intracellular cell organelle's structural alterations after photodynamic treatment with chlorin p6 -histamine conjugate (Cp6 -his) in human oral cancer cells. Herein, the cells were treated with Cp6 -his (10μm) and counterstained with organelle-specific fluorescence probes to find the site of intracellular localization using confocal microscopy. For photodynamic therapy (PDT), the cells were exposed to ~30 kJ/m2 red light (660 ± 20 nm) to induce ~90% cytotoxicity. We used the three-dimensional (3D) image reconstruction approach to analyze the photodynamic damage to cell organelles. The result showed that Cp6 -his localized mainly in the endoplasmic reticulum (ER) and lysosomes but not in mitochondria and Golgi apparatus (GA). The 3D model revealed that in necrotic cells, PDT led to extensive fragmentation of ER and fragmentation and swelling of GA as well. Results suggest that the indirect damage to GA occurred due to loss of connection between ER and GA. Moreover, in damaged cells with no sign of necrosis, the perinuclear ER appeared condensed and surrounded by several small clumps at the peripheral region of the cell, and the GA was observed to form a single condensed structure. Since these structural changes were associated with apoptotic cell death, it is suggested that the necrotic and apoptotic death induced by PDT with Cp6 -his is determined by the severity of damage to ER and indirect damage to GA. The results suggest that the indirect damage to cell organelle apart from the sites of photosensitizer localization and the severity of damage at the organelle level contribute significantly to the mode of cell death in PDT.

Family with sequence similarity 134 member B-mediated reticulophagy ameliorates hepatocyte apoptosis induced by dithiothreitol.

BACKGROUNDEndoplasmic reticulum (ER) stress-related hepatocyte apoptosis is responsible for multiple hepatic diseases. Previous studies have revealed that endoplasmic reticulophagy (ER-phagy) promotes the selective clearance of damaged ER fragments during ER stress, playing a crucial role in maintaining ER homeostasis and inhibiting apoptosis. Family with sequence similarity 134 member B (FAM134B) is a receptor involved in ER-phagy that can form a complex with calnexin (CNX) and microtubule-associated protein 1 light chain 3 (LC3). The complex can mediate the selective isolation of ER fragments to attenuate hepatocyte apoptosis. However, the precise regulatory mechanisms remain unclear.AIMTo elucidate the effect of FAM134B-mediated ER-phagy on ER stress-induced apoptosis in buffalo rat liver 3A (BRL-3A) rat hepatocytes and the potential regulatory mechanisms.METHODSER stress-related hepatocyte apoptosis was induced using dithiothreitol (DTT). Proteins related to ER stress and autophagy were measured with western blotting. Protein complex interactions with FAM134B were isolated by co-immunoprecipitation. ER-phagy was evaluated in immunofluorescence experiments. Cell cycle distribution and apoptosis were measured by flow cytometry. Mitochondrial Ca2+ levels were evaluated by the co-localization of intracellular Ca2+-tracker and Mito-tracker. The small interfering RNA against FAM134B was used to knockdown FAM134B in BRL-3A cells.RESULTSER stress-related and autophagy-related proteins in BRL-3A cells were elevated by both short and long-term DTT treatment. Furthermore, co-immunoprecipitation confirmed an interaction between FAM134B, CNX, FAM134B, and LC3 in BRL-3A cells. Immunofluorescence assays revealed that autolysosomes significantly decreased following short-term DTT treatment, but increased after long-term treatment. Mitochondrial Ca2+ levels and apoptotic rates were dramatically elevated, and more cells were arrested in the G1 stage after short-term DTT treatment; however, these decreased 48 h later. Moreover, FAM134B downregulation accelerated mitochondrial apoptotic pathway activation and aggravated hepatocyte apoptosis under ER stress.CONCLUSIONFAM134B-mediated ER-phagy attenuates hepatocyte apoptosis by suppressing the mitochondrial apoptotic pathway. Our findings provide new evidence highlighting the importance of FAM134B-mediated ER-phagy in attenuating hepatocyte apoptosis.

Doubly Stimulated Corrole for Organelle-Selective Antitumor Cytotoxicity.

Balancing between safety and efficacy of cancer chemotherapeutics is achievable by relying on internal and/or external stimuli for selective and on-demand antitumor cytotoxicity. We now introduce the difluorophosphorus(V) corrole PC-Im, a theranostic agent with a pH-sensitive N-methylimidazole moiety. Structure/activity relationships, via comparison with the permanently charged PC-ImM+ and the lipophilic PC, uncovered the exceptional features of PC-Im: nanoparticular and monomeric at neutral and low pH, respectively, 10-fold increased light-induced singlet oxygen production at acidic pH, internalization into malignant cells within minutes, and selective accumulation within lysosomes. Submillimolar PC-Im concentrations are tolerable in the dark, while illumination induces nanomolar cytotoxic effects due to a multiplicity of cellular deleterious events: endoplasmic reticulum fragmentation, lysosome fusion and exocytosis, calcium leakage, mitochondrial fission, and swelling. PC-Im emerges as an antitumor agent, whose potency is triggered by endogenous and exogenous stimuli, assuring its cytotoxicity will occur selectively upon lysosomal accumulation and solely upon light activation.

The unfolded protein response transducer IRE1α promotes reticulophagy in podocytes

Glomerular diseases involving podocyte/glomerular epithelial cell (GEC) injury feature protein misfolding and endoplasmic reticulum (ER) stress. Inositol-requiring enzyme 1α (IRE1α) mediates chaperone production and autophagy during ER stress. We examined the role of IRE1α in selective autophagy of the ER (reticulophagy). Control and IRE1α knockout (KO) GECs were incubated with tunicamycin to induce ER stress and subjected to proteomic analysis. This showed IRE1α-dependent upregulation of secretory pathway mediators, including the coat protein complex II component Sec23B. Tunicamycin enhanced expression of Sec23B and the reticulophagy adaptor reticulon-3-long (RTN3L) in control, but not IRE1α KO GECs. Knockdown of Sec23B reduced autophagosome formation in response to ER stress. Tunicamycin stimulated colocalization of autophagosomes with Sec23B and RTN3L in an IRE1α-dependent manner. Similarly, during ER stress, glomerular α5 collagen IV colocalized with RTN3L and autophagosomes. Degradation of RTN3L and collagen IV increased in response to tunicamycin, and the turnover was blocked by deletion of IRE1α; thus, the IRE1α pathway promotes RTN3L-mediated reticulophagy and collagen IV may be an IRE1α-dependent reticulophagy substrate. In experimental glomerulonephritis, expression of Sec23B, RTN3L, and LC3-II increased in glomeruli of control mice, but not in podocyte-specific IRE1α KO littermates. In conclusion, during ER stress, IRE1α redirects a subset of Sec23B-positive vesicles to deliver RTN3L-coated ER fragments to autophagosomes. Reticulophagy is a novel outcome of the IRE1α pathway in podocytes and may play a cytoprotective role in glomerular diseases.

ER-phagy: mechanisms, regulation, and diseases connected to the lysosomal clearance of the endoplasmic reticulum.

ER-phagy (reticulophagy) defines the degradation of portions of the endoplasmic reticulum (ER) within lysosomes or vacuoles. It is part of the self-digestion (i.e., autophagic) programs recycling cytoplasmic material and organelles, which rapidly mobilize metabolites in cells confronted with nutrient shortage. Moreover, selective clearance of ER subdomains participates in the control of ER size and activity during ER stress, the reestablishment of ER homeostasis after ER stress resolution, and the removal of ER parts in which aberrant and potentially cytotoxic material has been segregated. ER-phagy relies on the individual and/or concerted activation of the ER-phagy receptors, ER peripheral or integral membrane proteins that share the presence of LC3/Atg8-binding motifs in their cytosolic domains. ER-phagy involves the physical separation of portions of the ER from the bulk ER network and their delivery to the endolysosomal/vacuolar catabolic district. This last step is accomplished by a variety of mechanisms including macro-ER-phagy (in which ER fragments are sequestered by double-membrane autophagosomes that eventually fuse with lysosomes/vacuoles), micro-ER-phagy (in which ER fragments are directly engulfed by endosomes/lysosomes/vacuoles), or direct fusion of ER-derived vesicles with lysosomes/vacuoles. ER-phagy is dysfunctional in specific human diseases, and its regulators are subverted by pathogens, highlighting its crucial role for cell and organism life.

Anti-Inflammatory Anthranilate Analogue Enhances Autophagy through mTOR and Promotes ER-Turnover through TEX264 during Alzheimer-Associated Neuroinflammation.

Autophagy degrades impaired organelles and toxic proteins to maintain cellular homeostasis. Dysregulated autophagy is a pathogenic participant in Alzheimer's disease (AD) progression. In early-stage AD, autophagy is beneficially initiated by mild endoplasmic reticulum (ER) stress to alleviate cellular damage and inflammation. However, chronic overproduction of toxic Aβ oligomers eventually causes Ca2+ dysregulation in the ER, subsequently elevating ER-stress and impairing autophagy. Our previous work showed that a novel anthranilate analogue (SI-W052) inhibited lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-α and interleukin (IL)-6 on microglia. To investigate its mechanism of action, herein, we postulate that SI-W052 exhibits anti-inflammatory activity through ER-stress-mediated autophagy. We initially demonstrate that autophagy inhibits inflammation, but it becomes impaired during acute inflammation. SI-W052 significantly induces the conversion ratio of LC3 II/I and inhibits LPS-upregulated p-mTOR, thereby restoring impaired autophagy to modulate inflammation. Our signaling study further indicates that SI-W052 inhibits the upregulation of ER-stress marker genes, including Atf4 and sXbp1/tXbp1, explaining compound activity against IL-6. This evidence encouraged us to evaluate ER-stress-triggered ER-phagy using TEX264. ER-phagy mediates ER-turnover by the degradation of ER fragments to maintain homeostasis. TEX264 is an important ER-phagy receptor involved in ATF4-mediated ER-phagy under ER-stress. In our study, elevated TEX264 degradation is identified during inflammation; SI-W052 enhances TEX264 expression, producing a positive effect in ER-turnover. Our knockdown experiment further verifies the important role of TEX264 in SI-W052 activity against IL-6 and ER-stress. In conclusion, this study demonstrates that an anthranilate analogue is a novel neuroinflammation agent functioning through ER-stress-mediated autophagy and ER-phagy mechanisms.

Targeting FAM134B-mediated reticulophagy activates sorafenib-induced ferroptosis in hepatocellular carcinoma

Ferroptosis is a kind of cell death closely related to selective autophagy, such as ferritinophagy, lipophagy, clockophagy and chaperone-mediated autophagy. However, the role of reticulophagy, which specifically degrades endoplasmic reticulum (ER) fragments (also known as ER-phagy), in ferroptosis regulation is still unclear. In this study, we found that sorafenib (ferroptosis inducer) can effectively activate the receptor protein FAM134B-mediated ER-phagy, and FAM134B knockdown not only blocked ER-phagy but also significantly strengthened cellular sensitivity to ferroptosis without affecting macroautophagy. In vivo experiments also yielded similar results. These evidences provided new clues for ferroptosis regulation. Subsequently, bioinformatic analysis combined with RNA binding protein immunoprecipitation and polyribosome fractionation preliminarily indicated that PABPC1 can interact with FAM134B mRNA and promote its translation. Taken together, this study revealed the role of the PABPC1-FAM134B-ER-phagy pathway on ferroptosis, providing important evidence for novel anti-cancer strategies.

ER-Phagy and Microbial Infection.

The endoplasmic reticulum (ER) is an essential organelle in cells that synthesizes, folds and modifies membrane and secretory proteins. It has a crucial role in cell survival and growth, thus requiring strict control of its quality and homeostasis. Autophagy of the ER fragments, termed ER-phagy or reticulophagy, is an essential mechanism responsible for ER quality control. It transports stress-damaged ER fragments as cargo into the lysosome for degradation to eliminate unfolded or misfolded protein aggregates and membrane lipids. ER-phagy can also function as a host defense mechanism when pathogens infect cells, and its deficiency facilitates viral infection. This review briefly describes the process and regulatory mechanisms of ER-phagy, and its function in host anti-microbial defense during infection.