Lysosomal Autophagy Research Articles

Lysosome-related proteins may have changes in the urinary exosomes of patients with acute gout attack

BackgroundThe autophagy–lysosome is intricately linked to the development of gout. At present, the diagnosis and monitoring of gout are mainly invasive tests, which cannot predict the occurrence of gout in the acute phase, and bring new pain to patients. This study focuses on the changes of lysosome-related proteins in urinary exosomes of patients with acute gout attack to explore the potential noninvasive biomarkers clinical application value.MethodsUrine samples were collected from the subject and exosomes were extracted. To explore the differentially expressed proteins in urinary exosomes among acute gout patients (AD group), intermittent gout patients (ID group) and normal controls (NC group) by DIA mass spectrometry. Urinary exosomal lysosome associated proteins were analyzed and receiver operating characteristic (ROC) curves of differentially expressed proteins were drawn to evaluate their clinical value in monitoring acute gout attack.ResultsA total of 1896 proteins were detected between AD group and ID group, of which 121 proteins were differentially expressed (FC > 1.5 and p < 0.05). There were three lysosomal-related proteins differentially expressed in urinary exosomes between AD group and ID group. Compared with the ID group, the expression of Cathepsin Z (CTSZ) and AP-1 complex subunit beta-1 (AP1B1) was increased, while the expression of Lysosome-associated membrane glycoprotein 2 (LAMP2) was decreased in AD group. The ROC analysis showed that CTSZ, AP1B1 and LAMP2 had a strong ability to predict acute gout attack, with AUC of 0.826, 0.847 and 0.882, respectively.ConclusionsThere are many specific protein changes in the urinary exosomes of patients with acute gout attack. The urinary exosomes of patients with acute gout attack may exhibit alterations in lysosome-related proteins, particularly CTSZ, AP1B1, and LAMP2, which may become potential biomarkers for monitoring acute gout attack.

THSWD upregulates the LTF/AMPK/mTOR/Becn1 axis and promotes lysosomal autophagy in hepatocellular carcinoma cells by regulating gut flora and metabolic reprogramming.

THSWD has the effect of reducing inflammation, improving microcirculation, and regulating immune status in patients with hepatocellular carcinoma. Regardless of its clear therapeutic effect, the underlying mechanism of action against hepatocellular carcinoma is not clear. To identify critical gut microbiota and its associated metabolites related to THSWD inhibition against hepatocellular carcinoma progression, we assessed the microbe-dependent anti-hepatocellular carcinoma effects of THSWD through 16s rRNA gene sequencing, fecal microbial transplantation and antibiotic treatment. Metabolic analyses, transcriptomic analyses, and molecular experiments were performed to explore how THSWD modulates the gut microbiota against hepatocellular carcinoma progression. As confirmed by in vivo and in vitro assays, THSWD reduced tumour growth rate and promoted apoptosis in hepatocellular carcinoma cells in hepatocellular carcinoma model mice, and liver and kidney indexes were detected and confirmed the safety of THSWD. Transcriptomic analysis revealed that the targets of THSWD were significantly enriched in multiple lysosomal autophagy signalling pathways, suggesting that lysosomal autophagy is probably associated with THSWD's therapeutic effect. Based on the integrated data analysis, THSWD delays hepatocellular carcinoma progression by increasing the intestinal microbiota Duncaniella and augmenting the metabolite glabrol, and the joint analysis of metabolic and genomic data suggests that this metabolite is associated with lysosomal autophagy, and cellular experiments confirmed that the The differential metabolite glabrol induces apoptosis in hepatocellular carcinoma cells by triggering the lysosomal autophagy-mediated apoptosis signalling pathway. Supplementation with glabrol metabolites up regulates the LTF/AMPK/mTOR/Beclin1 axis and promotes hepatocellular carcinoma cells with lysosomal autophagy and induced apoptosis in hepatocellular carcinoma cells.

YTHDF3-mediated FLCN/cPLA2 axis improves cardiac fibrosis via suppressing lysosomal function.

Cardiac fibrosis characterized by aberrant activation of cardiac fibroblasts impairs cardiac contractile and diastolic functions, inducing the progression of the disease towards its terminal phase, resulting in the onset of heart failure. Therefore, the inhibition of cardiac fibrosis has become a promising treatment for cardiac diseases. The ovarian follicle-stimulating hormone folliculin (FLCN) plays a significant role in various biological processes, such as lysosome function, mitochondrial synthesis, angiogenesis, ciliogenesis and autophagy. Severe heart failure was observed in FLCN knockout mice. In this study, we investigated the role of FLCN in cardiac fibrosis and its potential mechanisms. The mice were subjected to transverse aortic constriction (TAC) surgery. Myocardial fibrosis developed in the mice 8 weeks after surgery. We showed that the protein and mRNA expression levels of FLCN were significantly decreased in TAC mice. Similar results were observed in primary mouse cardiac fibroblasts treated with Ang-II, an in vitro cardiac fibrosis model, suggesting that FLCN is involved in the pathological process of cardiac fibrosis. We demonstrated that overexpression of FLCN inhibited lysosome function in cardiac fibroblasts. Furthermore, overexpression of FLCN protected the heart from TAC-induced pathological cardiac fibrosis. We revealed that FLCN bound to the cPLA2 protein, increased its activity, regulated lysosomal function, and promoted membrane permeabilisation in cardiac fibroblasts during cardiac fibrosis. Knockdown of cPLA2 blocked the antifibrotic effect of FLCN in cardiac fibrosis. In addition, we found that the reduced expression of FLCN in cardiac fibrosis resulted from the modulation of YTHDF3-regulated m6A methylation of FLCN mRNA. The overexpression of YTHDF3 alleviated the production of collagens and improved cardiac structure and function in TAC mice. YTHDF3 inhibited proliferation and differentiation and regulated lysosomal function in mouse cardiac fibroblasts, whereas these effects were abolished by FLCN knockdown. We conclude that FLCN undergoes YTHDF3-regulated m6A modification and interacts with cPLA2 to improve lysosomal function in cardiac fibroblasts, highlighting its role in myocardial fibrosis therapy. These results suggest that FLCN and YTHDF3 could serve as potential therapeutic targets for cardiac fibroblast treatment.

Trehalose Ameliorates Zebrafish Emotional and Social Deficits Caused by CLN8 Dysfunction.

CLN8 and other neuronal ceroid lipofuscinoses (NCLs) often lead to cognitive decline, emotional disturbances, and social deficits, worsening with disease progression. Disrupted lysosomal pH, impaired autophagy, and defective dendritic arborization contribute to these symptoms. Using a cln8-/- zebrafish model, we identified significant impairments in locomotion, anxiety, and aggression, along with subtle deficits in social interactions, positioning zebrafish as a useful model for therapeutic studies in NCL. Our findings show that trehalose, an autophagy enhancer, ameliorates anxiety, and modestly improves social behavior and predator avoidance in mutant zebrafish. This finding aligns animal models with clinical reports suggestive of behavioral improvements in NCL patients. Trehalose holds promise as a therapeutic agent for CLN8, warranting further research into its neuroprotective mechanisms and clinical applications.

Nicotinamide mononucleotide protects septic hearts in mice via preventing cyclophilin F modification and lysosomal dysfunction.

Myocardial dysfunction is a decisive factor of death in septic patients. Cyclophilin F (PPIF) is a major component of the mitochondrial permeability transition pore (mPTP) and acts as a critical mPTP sensitizer triggering mPTP opening. In sepsis, decreased NAD+ impairs Sirtuin 3 function, which may prevent PPIF de-acetylation. Repletion of NAD+ with nicotinamide mononucleotide (NMN) reduces myocardial dysfunction in septic mice. In addition, administration of the mPTP inhibitor cyclosporine-A attenuated sepsis-induced myocardial dysfunction, and deletion of PPIF reduced lung and liver injuries in sepsis, leading to increased survival. It is plausible that NAD+ repletion with NMN may prevent mPTP opening in protecting septic hearts through PPIF de-acetylation and/or inhibition of mitochondrial ROS-mediated PPIF oxidation. In this study we investigated how NMN alleviated myocardial dysfunction in septic mice. Sepsis was induced in mice by injection of LPS (4 mg/kg, i.p.). Then mice received NMN (500 mg/kg, i.p.) or mito-TEMPO (0.7 mg/kg, i.p.) right after LPS injection, and subjected to echocardiography for assessing myocardial function. At the end of experiment, the heart tissues and sera were collected for analyses. In vitro experiments were conducted in neonatal mouse cardiomyocytes treated with LPS (1 µg/mL) in the presence of NMN (500 µmol/L) or mito-TEMPO (25 nmol/L). We showed that LPS treatment markedly increased mitochondrial ROS production and induced lysosomal dysfunction and aberrant autophagy in cardiomyocytes and mouse hearts, leading to inflammatory responses and myocardial injury and dysfunction in septic mice. NMN administration attenuated LPS-induced deteriorative effects. Selective inhibition of mitochondrial superoxide production with mito-TEMPO attenuated lysosomal dysfunction and aberrant autophagy in septic mouse hearts. Notably, LPS treatment significantly increased acetylation and oxidation of PPIF, which was prevented by NMN in mouse hearts. Knockdown of PPIF replicated the beneficial effects of NMN or mito-TEMPO on ROS production, lysosomal dysfunction, aberrant autophagy, and myocardial injury/dysfunction in sepsis. In addition, administration of NMN abrogated LPS-induced ATP5A1 acetylation and increased ATP5A1 protein levels and ATP production in septic mouse hearts. This study demonstrates that NMN modulates the interplay of mitochondrial ROS and PPIF in maintaining normal lysosomal function and autophagy and protecting ATP5A1 and ATP production during sepsis.

Autophagy-activating aluminum hydroxide nanovaccine for enhanced antigen presentation and anti-tumor immunity

Lymph node (LN) targeting and antigen presentation by antigen-presenting cells (APCs) are critical factors affecting the immune responses induced by tumor vaccines. Autophagy activation promotes MHC class I and II antigen presentation in APCs. To enhance antigen presentation in LNs, we developed an aluminum hydroxide nanovaccine that simultaneously incorporates the autophagy-activating peptide Beclin-1 and the antigenic protein OVA (B/O@AN nanovaccine) through layer-by-layer electrostatic interaction. B/O@AN has a particle size of approximately 80 nm and efficiently targets lymph nodes following subcutaneous administration. The combination of the Beclin-1 peptide with the aluminum hydroxide nanovaccine promotes dendritic cell (DC) maturation. More importantly, B/O@AN facilitates antigen cross-presentation by promoting lysosomal escape and autophagy induction. After immunization, compared to O/@AN without Beclin-1, B/O@AN significantly augments antigen-specific cellular immune responses, leading to substantial increases in cytotoxic T lymphocytes (CTLs), T-helper 1 (Th1) cells, as well as serum antibody levels, thereby impeding melanoma development and progression in both prophylactic and therapeutic settings. These results provide evidence that autophagy activation strengthens antigen presentation and augments the antigen-specific immune responses of the aluminum hydroxide nanovaccine.

Adverse outcome pathway of Alzheimer's disease-like changes resulting from autophagy flux blockade after MC-LR exposure

Microcystins (MCs) pollution is a worldwide environmental issue concerning about human health. Microcystin-leucine-arginine (MC-LR), the most common type of MCs produced by cyanobacteria, could enter the brain and bring about damage to the nervous system. Up to date, it is not clear about the mechanism of MC-LR-induced neurotoxicity. Amyloid-β (Aβ) deposits are hallmark of Alzheimer's disease (AD). In this study, we revealed that MC-LR exposure at environment-related doses (1, 7.5, 15 μg/L) could promote Aβ accumulation in mouse brain. Mechanically, we firstly found that Aβ accumulation is closely associated with abnormal Aβ degradation due to autophagy flux blockade and lysosome dysfunctions in neurons after MC-LR exposure. Moreover, an adverse outcome pathway (AOP) framework oriented to neurotoxicity of MC-LR was conducted in this study. MC-LR inhibited the activity of protein phosphatase 2A (PP2A) in neurons, which is regarded as a molecular initiating event (MIE). In addition, the abnormalities in autophagy were observed after MC-LR exposure. The hindered autophagosome-lysosome fusion and disrupted lysosomal function were key events (KEs) after MC-LR exposure, which contributed to proteostasis dysregulation, ultimately leading to Aβ abnormal degradation and learning deficits as adverse outcomes (AO) of neurotoxicity. This study provided novel information about MC-LR neurotoxicity and new insights into understanding the mechanisms underlying the environmental chemicals-induced neurodegeneration diseases, which has deep implications for public health.

Schaftoside improves HFpEF through regulation the autophagy-lysosome pathway by allosterically targeting CaMKII-δ

Heart failure with preserved ejection fraction (HFpEF) presents a significant challenge to global healthcare systems due to its complex presentation. HFpEF presents with a normal or near-normal left ventricular ejection fraction, cardiac diastolic dysfunction, and a metabolic profile characterized by impaired inflammation and oxidative stress. There have been few valuable drug targets reported for HFpEF to date. Here, we discovered that schaftoside, an active component from licorice, has a significant protective effect on the cardiac remodeling induced by continuous infusion of angiotensin II (AngII), which leads to the HFpEF phenotype. Mechanistically, schaftoside has demonstrated the ability to ameliorate lysosomal dysfunction in both in vitro and in vivo models, thereby activating autophagy. Bioinformatic analyses based on proteome and phosphoproteome suggested that Ca2+/calmodulin-dependent protein kinase II (CaMKII) was a potential target for schaftoside. It was confirmed that schaftoside allosterically mediated CaMKII-δ conformation via targeting a unique active pocket near the ATP-binding site to inhibit protein phosphorylation and regulate the lysosomal autophagy pathway. Therefore, schaftoside represents the first small molecule identified to inhibit CaMKII-δ activity through allosteric inhibition, providing a novel candidate for alleviating cardiac metabolic imbalance in HFpEF.

TMEM175 plays a crucial role in osteoblast differentiation by regulating lysosomal function and autophagy

Bone provides structural support, enables movement, protects internal organs, regulates calcium and phosphorus levels, and contains bone marrow essential for hematopoiesis. Osteoblasts are specialized cells responsible for bone formation through the secretion of extracellular matrix components. Transmembrane protein 175 (TMEM175), which functions as an endosomal/lysosomal K+ channel and a lysosomal H+ channel, regulates lysosomal function and autophagy. Despite the recognized importance of lysosomes and autophagy in osteoblast differentiation, the specific role of TMEM175 in osteoblast differentiation has not been revealed. In this study, we investigated whether TMEM175 is associated with human bone mineral densityand fracture and examined the role of TMEM175 in osteoblast differentiation. In analyses of single nucleotide polymorphismsof pore ion channel genes using the mouse2human database, a significant correlation between TMEM175 single nucleotide polymorphisms and human bone mineral density and fracture was identified. TMEM175 expression levels were found to increase during osteoblast differentiation from bone chip-derived mesenchymal stem cells (BMSCs). Knockdown of TMEM175 in BMSCs suppressed osteoblast differentiation, as evidenced by decreased matrix mineralization and lower expression levels of osteoblast marker genes. Further analysis indicated that TMEM175 deficiency leads to lysosomal dysfunction and partially impairs autophagic clearance during osteoblast differentiation. Moreover, the TMEM175 inhibitor 4-aminopyridinedecreased osteoblast differentiation of BMSCs. Taken together, this study reveals that TMEM175 plays an important role in osteoblast differentiation by regulating lysosomal function and autophagic clearance.

Jiedu recipe, a compound Chinese herbal medicine, suppresses hepatocellular carcinoma metastasis by inhibiting the release of tumor-derived exosomes in a hypoxic microenvironment

ObjectiveTumor-derived exosomes (TDEs) play crucial roles in intercellular communication. Hypoxia in the tumor microenvironment enhances secretion of TDEs and accelerates tumor metastasis. Jiedu recipe (JR), a traditional Chinese medicinal formula, has demonstrated efficacy in preventing the metastasis of hepatocellular carcinoma (HCC). However, the underlying mechanism remains largely unknown. MethodsAnimal experiments were performed to investigate the metastasis-preventing effects of JR. Bioinformatics analysis and in vitro assays were conducted to explore the potential targets and active components of JR. TDEs were assessed using nanoparticle tracking analysis (NTA) and Western blotting (WB). Exosomes derived from normoxic or hypoxic HCC cells (H-TDEs) were collected to establish premetastatic mouse models. JR was intragastrically administered to evaluate its metastasis-preventive effects. WB and lysosomal staining were performed to investigate the effects of JR on lysosomal function and autophagy. Bioinformatics analysis, WB, NTA, and immunofluorescence staining were used to identify the active components and potential targets of JR. ResultsJR effectively inhibited subcutaneous-tumor-promoted lung premetastatic niche development and tumor metastasis. It inhibited the release of exosomes from tumor cells under hypoxic condition. JR treatment promoted both lysosomal acidification and suppressed secretory autophagy, which were dysregulated in hypoxic tumor cells. Quercetin was identified as the active component in JR, and the epidermal growth factor receptor (EGFR) was identified as a potential target. Quercetin inhibited EGFR phosphorylation and promoted the nuclear translocation of transcription factor EB (TFEB). Hypoxia-impaired lysosomal function was restored, and secretory autophagy was alleviated by quercetin treatment. ConclusionJR suppressed HCC metastasis by inhibiting hypoxia-stimulated exosome release, restoring lysosomal function, and suppressing secretory autophagy. Quercetin acted as a key component of JR and regulated TDE release through EGFR-TFEB signaling. Our study provides a potential strategy for retarding tumor metastasis by targeting H-TDE secretion.Please cite this article as: Jia WT, Xiang S, Zhang JB, Yuan JY, Wang YQ, Liang SF, Lin WF, Zhai XF, Shang Y, Ling CQ, Cheng BB. Jiedu recipe, a compound Chinese herbal medicine, suppresses hepatocellular carcinoma metastasis by inhibiting the release of tumor-derived exosomes in a hypoxic microenvironment through the EGFR-TFEB signaling pathway. J Integr Med. 2024; Epub ahead of print.

Phospholipase D3 regulates TFEB/TFE3 metabolism to maintain lysosomal homeostasis.

A coding variant in Phospholipase D3 ( PLD3 ) increases the risk of Alzheimer's disease (AD). PLD3 is a lysosomal protein, and endosomal and lysosomal abnormalities are linked to AD; however, the role of PLD3 in lysosomal homeostasis and its implications in AD remain poorly understood. To address this knowledge gap, we conducted comprehensive studies integrating transcriptomics, proteomics, and cell biology approaches. We observed significant enlargement of lysosomes in neurons lacking PLD3, accompanied by increased endocytosis and autophagy, but a decline in lysosomal proteolytic activity. Lysosomes of PLD3-deficient cells underwent proteome remodeling, manifested by an enrichment of proteins involved in lysosomal biogenesis, endocytosis and calcium signaling. Mechanistically, we discovered that PLD3 mediates TFEB/TFE3 degradation through the proteasome, and as a result, PLD3 deficiency leads to increased TFEB/TFE3 levels, nuclear translocation, and transcriptional activities. Notably, variants in PLD3, e.g., V232M or K486R, do not alter its impact on TFEB/TFE3 metabolism. Transcriptomic profiling further confirmed the enrichment of transcripts involved in lysosomal biogenesis, endocytosis, autophagy, mTOR signaling and AD in response to PLD3 loss. Additionally, PLD3 ablation has synergistic effects with β-amyloid in causing lysosomal abnormalities and modifying TFEB/TFE3 signaling. In conclusion, our findings demonstrate that PLD3 is involved in regulating lysosomal biogenesis via TFEB/TFE3 signaling, and lysosomal abnormalities resulting from PLD3 deficiency are potentially a risk factor for AD.

Autophagic inhibitor ROC-325 ameliorates glomerulosclerosis and podocyte injury via inhibiting autophagic flux in experimental FSGS mice

BackgroundAutophagy plays an important role in the pathogenesis of focal segmental glomerulosclerosis (FSGS). Podocyte-specific Yes-associated protein (YAP) deletion mice, referred to as YAP-KO mice, is considered a new animal model to study the underlying mechanism of FSGS. ROC-325 is a novel small-molecule lysosomal autophagy inhibitor that is more effective than chloroquine (CQ) and hydroxychloroquine (HCQ) in suppressing autophagy. In this study, we sought to determine the therapeutic benefit and mechanism of action of ROC-325 in YAP-KO mice, an experimental FSGS model. Methods and resultsYAP-KO mice were treated with ROC-325 (50 mg/kg, p.o.) daily for one month. Our results revealed that albuminuria, mesangial matrix expension, and focal segmental glomerulosclerosis in YAP-KO mice were significantly attenuated by ROC-325 administration. Transmission electron microscopy and immunofluorescence staining showed that ROC-325 treatment significantly inhibited YAP-KO-induced autophagy activation by decreasing autophagosome-lysosome fusion and increasing LC3A/B and p62/SQSTM. Meanwhile, Immunofluorescence staining revealed that preapplication of ROC-325 in podocyte with YAP-targeted siRNA and mRFP-GFP-LC3 adenovirus markedly suppressed autophagic flux in vitro, suggesting that autophagy intervention may serve as a target for FSGS. ConclusionsThese results showed that the role of autophagic activity in FSGS mice model and ROC-325 could be a novel and promising agent for the treatment of FSGS.

A TBK1-independent primordial function of STING in lysosomal biogenesis

Stimulator of interferon genes (STING) is activated in many pathophysiological conditions, leading to TBK1-dependent interferon production in higher organisms. However, primordial functions of STING independent of TBK1 are poorly understood. Here, through proteomics and bioinformatics approaches, we identify lysosomal biogenesis as an unexpected function of STING. Transcription factor EB (TFEB), an evolutionarily conserved regulator of lysosomal biogenesis and host defense, is activated by STING from multiple species, including humans, mice, and frogs. STING-mediated TFEB activation is independent of TBK1, but it requires STING trafficking and its conserved proton channel. GABARAP lipidation, stimulated by the channel of STING, is key for STING-dependent TFEB activation. STING stimulates global upregulation of TFEB-target genes, mediating lysosomal biogenesis and autophagy. TFEB supports cell survival during chronic sterile STING activation, a common condition in aging and age-related diseases. These results reveal a primordial function of STING in the biogenesis of lysosomes, essential organelles in immunity and cellular stress resistance.

Autophagy-lysosomal-associated neuronal death in neurodegenerative disease.

Autophagy, the major lysosomal pathway for degrading damaged or obsolete constituents, protects neurons by eliminating toxic organelles and peptides, restoring nutrient and energy homeostasis, and inhibiting apoptosis. These functions are especially vital in neurons, which are postmitotic and must survive for many decades while confronting mounting challenges of cell aging. Autophagy failure, especially related to the declining lysosomal ("phagy") functions, heightens the neuron's vulnerability to genetic and environmental factors underlying Alzheimer's disease (AD) and other late-age onset neurodegenerative diseases. Components of the global autophagy-lysosomal pathway and the closely integrated endolysosomal system are increasingly implicated as primary targets of these disorders. In AD, an imbalance between heightened autophagy induction and diminished lysosomal function in highly vulnerable pyramidal neuron populations yields an intracellular lysosomal build-up of undegraded substrates, including APP-βCTF, an inhibitor of lysosomal acidification, and membrane-damaging Aβ peptide. In the most compromised of these neurons, β-amyloid accumulates intraneuronally in plaque-like aggregates that become extracellular senile plaques when these neurons die, reflecting an "inside-out" origin of amyloid plaques seen in human AD brain and in mouse models of AD pathology. In this review, the author describes the importance of lysosomal-dependent neuronal cell death in AD associated with uniquely extreme autophagy pathology (PANTHOS) which is described as triggered by lysosomal membrane permeability during the earliest "intraneuronal" stage of AD. Effectors of other cell death cascades, notably calcium-activated calpains and protein kinases, contribute to lysosomal injury that induces leakage of cathepsins and activation of additional death cascades. Subsequent events in AD, such as microglial invasion and neuroinflammation, induce further cytotoxicity. In major neurodegenerative disease models, neuronal death and ensuing neuropathologies are substantially remediable by reversing underlying primary lysosomal deficits, thus implicating lysosomal failure and autophagy dysfunction as primary triggers of lysosomal-dependent cell death and AD pathogenesis and as promising therapeutic targets.

Lactylation stabilizes TFEB to elevate autophagy and lysosomal activity.

The transcription factor TFEB is a major regulator of lysosomal biogenesis and autophagy. There is growing evidence that posttranslational modifications play a crucial role in regulating TFEB activity. Here, we show that lactate molecules can covalently modify TFEB, leading to its lactylation and stabilization. Mechanically, lactylation at K91 prevents TFEB from interacting with E3 ubiquitin ligase WWP2, thereby inhibiting TFEB ubiquitination and proteasome degradation, resulting in increased TFEB activity and autophagy flux. Using a specific antibody against lactylated K91, enhanced TFEB lactylation was observed in clinical human pancreatic cancer samples. Our results suggest that lactylation is a novel mode of TFEB regulation and that lactylation of TFEB may be associated with high levels of autophagy in rapidly proliferating cells, such as cancer cells.

Premature senescence is regulated by crosstalk among TFEB, the autophagy lysosomal pathway and ROS derived from damaged mitochondria in NaAsO2-exposed auditory cells

Age-related hearing loss (ARHL) is one of the most prevalent types of sensory decline in a superaging society. Although various studies have focused on the effect of oxidative stress on the inner ear as an inducer of ARHL, there are no effective preventive approaches for ARHL. Recent studies have suggested that oxidative stress-induced DNA damage responses (oxidative DDRs) drive cochlear cell senescence and contribute to accelerated ARHL, and autophagy could function as a defense mechanism against cellular senescence in auditory cells. However, the underlying mechanism remains unclear. Sodium arsenite (NaAsO2) is a unique oxidative stress inducer associated with reactive oxygen species (ROS) that causes high-tone hearing loss similar to ARHL. Transcription factor EB (TFEB) functions as a master regulator of the autophagy‒lysosome pathway (ALP), which is a potential target during aging and the pathogenesis of various age-related diseases. Here, we focused on the function of TFEB and the impact of intracellular ROS as a potential target for ARHL treatment in a NaAsO2-induced auditory premature senescence model. Our results suggested that short exposure to NaAsO2 leads to DNA damage, lysosomal damage and mitochondrial damage in auditory cells, triggering temporary signals for TFEB transport into the nucleus and, as a result, causing insufficient autophagic flux and declines in lysosomal function and biogenesis and mitochondrial quality. Then, intracellular ROS derived from damaged mitochondria play a role as a second messenger to induce premature senescence in auditory cells. These findings suggest that TFEB activation via transport into the nucleus contributes to anti-senescence activity in auditory cells and represents a new therapeutic target for ARHL. We have revealed the potential function of TFEB as a master regulator of the induction of oxidative stress-induced premature senescence and the senescence-associated secretion phenotype (SASP) in auditory cells, which regulates ALP and controls mitochondrial quality through ROS production.

Nanohybrid-Based Redox Homeostasis Perturbators Escaped from Early Lysosomes toward Amplified Sensitization of Tumor Cells and Photothermally Maneuvered Pyroptosis Therapy.

Reactive oxygen species (ROS) hold great potential in tumor pyroptosis therapy, yet they are still limited by short species lifespan and limited diffusion distance. Inducing cells into a metastable state and then applying external energy can effectively trigger pyroptosis, but systemic sensitization still faces challenges, such as limited ROS content, rapid decay, and short treatment windows. Herein, a nanohybrid-based redox homeostasis-perturbator system was designed that synergistically induce early lysosomal escape, autophagy inhibition, and redox perturbation functions to effectively sensitize cells to address these challenges. Specifically, weakly alkaline layered double hydroxide nanosheets (LDH NSs) with pH-responsive degradation properties enabled early lysosomal escape within 4 h, releasing poly(L-dopa) nanoparticles for inducing catechol-quinone redox cycling in the cytoplasm. The intracellular ROS levels were systematically rebounded by 3-4 times in tumor cells and lasted for over 4 h. Subsequently induced lysosomal stress and Ca2+ signaling activation resulted in severe mitochondrial dysfunction, as well as a perilous metastable state. Thereby, sequential near-infrared light was applied to trigger amplified stress through a local photothermal conversion. This led to sufficiently high levels of cleaved caspase-1 and GSDMD activation (2.5-2.8-fold increment) and subsequent pyroptosis response. In addition, OH- released by LDH elevated pH to alleviate the limitation of glutathione depletion by quinones at acidic pH and inhibit protective autophagy. Largely secreted inflammatory factors (2.5-5.6-fold increment), efficient maturation of dendritic cells, and further immune stimulation were boosted for tumor inhibition as a consequence. This study offers a new paradigm and insights into the synergy of internal systematic cellular sensitization and sequential external energy treatment to achieve tumor suppression through pyroptosis.

PDCD4 triggers α-synuclein accumulation and motor deficits via co-suppressing TFE3 and TFEB translation in a model of Parkinson’s disease

TFE3 and TFEB, as the master regulators of lysosome biogenesis and autophagy, are well characterized to enhance the synaptic protein α-synuclein degradation in protecting against Parkinson’s disease (PD) and their levels are significantly decreased in the brain of PD patients. However, how TFE3 and TFEB are regulated during PD pathogenesis remains largely vague. Herein, we identified that programmed cell death 4 (PDCD4) promoted pathologic α-synuclein accumulation to facilitate PD development via suppressing both TFE3 and TFEB translation. Conversely, PDCD4 deficiency significantly augmented global and nuclear TFE3 and TFEB distributions to alleviate neurodegeneration in a mouse model of PD with overexpressing α-synuclein in the striatum. Mechanistically, like TFEB as we reported before, PDCD4 also suppressed TFE3 translation, rather than influencing its transcription and protein stability, to restrain its nuclear translocation and lysosomal functions, eventually leading to α-synuclein aggregation. We proved that the two MA3 domains of PDCD4 mediated the translational suppression of TFE3 through binding to its 5’-UTR of mRNA in an eIF-4A dependent manner. Based on this, we developed a blood-brain barrier penetrating RVG polypeptide modified small RNA drug against pdcd4 to efficiently prevent α-synuclein neurodegeneration in improving PD symptoms by intraperitoneal injections. Together, we suggest PDCD4 as a PD-risk protein to facilitate α-synuclein neurodegeneration via suppressing TFE3 and TFEB translation and further provide a potential small RNA drug against pdcd4 to treat PD by intraperitoneal injections.

An old dog with new tricks: TFEB promotes syncytin expression and cell fusion in the human placenta.

In the human placenta, cell fusion is crucial for forming the syncytiotrophoblast, a multinucleated giant cell essential for maintaining pregnancy and ensuring fetal health. The formation of the syncytiotrophoblast is catalyzed by the evolutionarily modern fusogens syncytin-1 and syncytin-2. In this issue of Genes & Development, Esbin and colleagues (doi:10.1101/gad.351633.124) reveal a critical role for the transcription factor TFEB in the regulation of syncytin expression and the promotion of trophoblast fusion. Notably, TFEB's pro-fusion role operates independently of its well-known functions in lysosome biogenesis and autophagy, suggesting that TFEB has acquired additional functions to promote cell fusion in the human placenta.

Cholesterol-rich lysosomes induced by respiratory syncytial virus promote viral replication by blocking autophagy flux

Respiratory syncytial virus (RSV) hijacks cholesterol or autophagy pathways to facilitate optimal replication. However, our understanding of the associated molecular mechanisms remains limited. Here, we show that RSV infection blocks cholesterol transport from lysosomes to the endoplasmic reticulum by downregulating the activity of lysosomal acid lipase, activates the SREBP2–LDLR axis, and promotes uptake and accumulation of exogenous cholesterol in lysosomes. High cholesterol levels impair the VAP-A-binding activity of ORP1L and promote the recruitment of dynein–dynactin, PLEKHM1, or HOPS VPS39 to Rab7–RILP, thereby facilitating minus-end transport of autophagosomes and autolysosome formation. Acidification inhibition and dysfunction of cholesterol-rich lysosomes impair autophagy flux by inhibiting autolysosome degradation, which promotes the accumulation of RSV fusion protein. RSV-F storage is nearly abolished after cholesterol depletion or knockdown of LDLR. Most importantly, the knockout of LDLR effectively inhibits RSV infection in vivo. These findings elucidate the molecular mechanism of how RSV co-regulates lysosomal cholesterol reprogramming and autophagy and reveal LDLR as a novel target for anti-RSV drug development.