Autophagy最新文献

The endosomal sorting complex required for transport (ESCRT) machinery is a membrane abscission system that mediates various intracellular membrane remodeling processes, including macroautophagy/autophagy. In our recent study, we established the unique requirement of the ubiquitin E2 variant-like (UEVL) domain of the ESCRT-I subunit VPS37A for phagophore closure, the final step in autophagosome biogenesis, and determined the physiological impact of systemically inhibiting closure by targeting this region in mice. While the mutant mice exhibited phenotypes similar to those reported in mice deficient in generating ATG8 (mammalian Atg8 homologs)-conjugated (ATG8ylated) phagophores, certain phenotypes, such as neonatal lethality and liver injury, were found to be notably milder. Further investigation revealed that ATG8ylated phagophores promote TBK1-dependent SQSTM1 phosphorylation and droplet formation, leading to the formation of large insoluble aggregates upon closure inhibition. These findings suggest potential roles for ATG8ylated membranes in mitigating proteotoxicity by efficiently concentrating and sequestering soluble, reactive microaggregates and converting them into less reactive, insoluble large aggregates. The study highlights VPS37A UEVL mutant mice as a model for investigating the physiological and pathological roles of phagophores that extend beyond degradation.

Myxomatous mitral valve degeneration (MMVD) is one of the most important age-dependent degenerative heart valve disorders in both humans and dogs. It is characterized by the aberrant remodeling of extracellular matrix (ECM), regulated by senescent myofibroblasts (aVICs) transitioning from quiescent valve interstitial cells (qVICs), primarily under TGFB1/TGF-β1 control. In the present study, we found senescent aVICs exhibited impaired macroautophagy/autophagy as evidenced by compromised autophagy flux and immature autophagosomes. MTOR-dependent autophagy induced by rapamycin and torin-1 attenuated cell senescence and decreased the expression of cyclin-dependent kinase inhibitors (CDKIs) CDKN2A/p16^INK4A and CDKN1A/p21^CIP1. Furthermore, induction of autophagy in aVICs by ATG (autophagy related) gene overexpression restored autophagy flux, with a concomitant reduction in CDKN1A and CDKN2A expression and senescence-associated secretory phenotype (SASP). Conversely, autophagy deficiency induced CDKN1A and CDKN2A accumulation and SASP, whereas ATG re-expression alleviated senescent phenotypic transformation. Notably, CDKN1A and CDKN2A localized to autophagosomes and lysosomes following MTOR antagonism or MG132 treatment. SQSTM1/p62 was identified as the autophagy receptor to selectively sequester CDKN1A and CDKN2A cargoes for autophagic degradation. Our findings are the first demonstration that CDKN1A and CDKN2A are degraded through SQSTM1-mediated selective autophagy, independent of the ubiquitin-proteasome pathway. These data will inform development of therapeutic strategies for the treatment of canine and human MMVD, and for the treatment of Alzheimer disease, Parkinson disease and other age-related degenerative disorders.

RAS mutations enhance macroautophagy/autophagy in tumor cells, crucial for their growth and survival, making autophagy a promising therapeutic target for RAS-mutant cancers. However, the distinction between RAS-induced autophagy and physiological autophagy is not well understood. We recently identified a unique form of autophagy, RAS-induced non-canonical autophagy via ATG8ylation (RINCAA), which differs from starvation-induced autophagy. RINCAA is regulated by different sets of autophagic factors and forms structures distinct from the double-membrane autophagosome known as RAS-induced multivesicular/multilaminar bodies of ATG8ylation (RIMMBA). A key feature of RINCAA is the phosphorylation of PI4KB by ULK1, and inhibiting this phosphorylation shows superior effects compared to general autophagy inhibitors. This work suggests a potential for specifically targeting autophagy in RAS-driven cancers as a therapeutic strategy.

The endoplasmic reticulum (ER) is a central hub for lipid metabolism and protein secretion, crucial for maintaining cellular homeostasis and mediating environmental interactions. ER-resident proteins VMP1 and TMEM41B function as scramblases, regulating lipid membranes to support macroautophagy and lipid droplet metabolism. To explore their developmental roles, we generated Vmp1 and Tmem41b mutations in mouse embryonic stem cells (ESCs). While these mutations did not affect ESC self-renewal or pluripotency, they impaired differentiation into the primitive endoderm lineage. Our findings reveal that this defect stems from VMP1 and TMEM41B's critical role in the maturation and stability of FZD2/FRIZZLED2, essential for WNT signaling. Thus, this study highlights their extensive role in protein trafficking, linking lipid metabolism to cell signaling and deepening our understanding of their diverse contributions to cellular and developmental processes.

Macroautophagy/autophagy is an evolutionarily conserved intracellular degradation pathway that relies on vacuoles or lysosomes. Over 40 ATG genes have been identified in yeast cells as participants in various types of autophagy, although these genes are non-essential. While some essential genes involved in autophagy have been identified using temperature-sensitive yeast strains, systematic research on essential genes in autophagy remains lacking. To address this, we established an essential protein conditional degradation library using the auxin-inducible degron (AID) system. By introducing the GFP-Atg8 plasmid, we identified 29 essential yeast genes involved in autophagy, 19 of which had not been previously recognized. In summary, the yeast essential protein conditional degradation library we constructed will serve as a valuable resource for systematically investigating the roles of essential genes in autophagy and other biological functions.

Ferroptosis is an iron-dependent regulated form of cell death implicated in various diseases, including cancers, with its progression influenced by iron-dependent peroxidation of phospholipids and dysregulation of the redox system. Whereas the extracellular matrix of tumors provides mechanical cues influencing tumor initiation and progression, its impact on ferroptosis and its mechanisms remains largely unexplored. In this study, we reveal that heightened mechanical tension sensitizes cells to ferroptosis, whereas decreased mechanics confers resistance. Mechanistically, reduced mechanical tension reduces intracellular free iron levels by enhancing FTH1 protein expression. Additionally, low mechanics significantly diminishes NCOA4, pivotal in mediating FTH1 phase separation-induced ferritinophagy. Targeting NCOA4 effectively rescues ferroptosis susceptibility under low mechanical tension through modulation of FTH1 phase separation-driven autophagy. In conclusion, our findings demonstrate that mechanics regulates iron metabolism via NCOA4-FTH1 phase separation-mediated autophagy, thereby influencing ferroptosis sensitivity and offering promising therapeutic avenues for future exploration.

Lysosomes are best known for their involvement in inflammatory responses, where they participate in the macroautophagy/autophagy process to eliminate inflammasomes. Recently, we have identified a previously overlooked function of lysosomes in regulating macrophage inflammatory responses. Specifically, lysosomes finely control the production of IL1B (interleukin 1 beta) by manipulating the release of lysosomal Fe²⁺ through MCOLN1. Mechanistically, reactive oxygen species (ROS), accumulated during sustained inflammation in macrophages, cause activation of the MCOLN1, a lysosomal cationic channel. The activation of MCOLN1 triggers the release of lysosomal Fe² toward the cytosol, which in turn activates prolyl hydroxylase domain enzymes (PHDs). PHDs' activation represses the transcriptional regulator NFKB/NF-kB (nuclear factor kappa B) activity by restraining RELA/p65 in the cytosol, leading to decreased IL1B transcription in macrophages. Consequently, the property of controlling production and subsequent release of IL1B from macrophages allows the lysosome to finely restrict sustained inflammatory responses. These findings demonstrate that apart from relying on its degradative capability, the lysosome also limits excessive inflammatory responses to facilitate the restoration of cellular and tissue homeostasis in macrophages by modulating the release of lysosomal Fe²⁺ through MCOLN1. Even more, by suppressing IL1B production, in vivo stimulation of the MCOLN1 channel alleviates multiple clinical symptoms of dextran sulfate sodium (DSS)-induced colitis in mice, highlighting MCOLN1 as a promising therapeutic target for inflammatory bowel disease (IBD) in clinical settings.

Stress granules (SGs) are transient, non-membrane-bound cytoplasmic condensates that form in response to environmental stresses, serving as protective reservoirs for mRNAs and proteins. In plants, SGs play a crucial role in stress adaptation, but their relationship with macroautophagy/autophagy, a key process for degrading damaged organelles and misfolded proteins, remains poorly understood. In a recent study, we revealed that key autophagy proteins, including components of the ATG1-ATG13 kinase complex, the class III phosphatidylinositol 3-kinase (PtdIns3K) complex, and the ATG8-PE system, translocate to SGs during heat stress (HS) in Arabidopsis thaliana. Using biochemical, cell biological and genetic approaches, we demonstrated that ATG proteins accumulate on HS-induced SGs and are released to the cytosol upon SG disassembly during the post-HS recovery stage. This process facilitates rapid autophagy activation. Notably, a SG-deficient mutant (ubp1abc) exhibits delayed autophagy activation and impaired clearance of ubiquitinated protein aggregates, highlighting the importance of SGs in regulating autophagy. Our findings uncover a novel mechanism by which SGs sequester autophagy proteins during stress, ensuring their rapid availability for stress recovery, and provide new insights into the interplay between SGs and autophagy in plant stress responses.Abbreviation: ATG, autophagy related; HS, heat stress; PtdIns3K, phosphatidylinositol 3-kinase; RBP47B, RNA-binding protein 47B; SG, stress granule; UBP1, ubiquitin-specific protease 1.

Protein translation is an energy-intensive ribosome-driven process that is reduced during nutrient scarcity to conserve cellular resources. During prolonged starvation, cells selectively translate specific proteins to enhance their survival (adaptive translation); however, this process is poorly understood. Accordingly, we analyzed protein translation and mRNA transcription by multiple methods in vitro and in vivo to investigate adaptive hepatic translation during starvation. While acute starvation suppressed protein translation in general, proteomic analysis showed that prolonged starvation selectively induced translation of lysosome and autolysosome proteins. Significantly, the expression of the orphan nuclear receptor, ESRRA (estrogen related receptor, alpha) increased during prolonged starvation and served as a master regulator of this adaptive translation by transcriptionally stimulating Rplp1 (ribosomal protein lateral stalk subunit P1) gene expression. Overexpression or siRNA knockdown of Esrra in vitro or in vivo led to parallel changes in Rplp1 gene expression, lysosome and macroautophagy/autophagy protein translation, and autophagy activity. Remarkably, we have found that ESRRA had dual functions by not only regulating transcription but also controlling adaptive translation via the ESRRA-RPLP1-lysosome-autophagy pathway during prolonged starvation.

Despite decades of research on effective methods to resist Salmonella enterica serovar Typhimurium (S. Typhimurium) pathogenicity, the mechanisms of S. Typhimurium-host interactions have not been fully determined. S. Typhimurium is characterized as an important zoonosis in public health worldwide because of its endemicity, high morbidity, and difficulty in applying control and prevention measures. Herein, we introduce a novel bacterial factor, secretion system effector J (SseJ), and its interactive host protein, PHB2 (prohibitin 2). We explored whether SseJ affected S. Typhimurium replication and survival in the host. S. Typhimurium infection caused severe mitochondrial damage and mitophagy, which facilitated S. Typhimurium proliferation in cells. S. Typhimurium SseJ activated the PINK1 (PTEN induced kinase 1)-PRKN (parkin RBR E3 ubiquitin protein ligase)-autophagosome-dependent mitophagy pathway, aided by the mitophagy receptor PHB2, for bacterial survival and persistent infection. Moreover, suppression of mitophagy alleviated the pathogenicity of S. Typhimurium. In conclusion, S. Typhimurium infection could be antagonized by targeting the SseJ-PHB2-mediated host mitochondrial autophagy pathway.Abbreviation: ACTB: actin beta; BafA1: bafilomycin A₁; CCCP: carbonyl cyanide m-chlorophenyl hydrazone; co-IP: co-immunoprecipitation; CFU: colony-forming units; COX4/COXIV: cytochrome c oxidase subunit 4; CQ: chloroquine; hpi: h post-bacterial infection; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; Mdivi-1:mitophagy inhibitor mitochondrial division inhibitor 1; MFN2: mitofusin 2; MG132: z-leu-leu-leucinal; MOI: multiplicity of infection; mtDNA: mitochondrial DNA; PBS: phosphate-buffered saline; PGAM5: PGAM family member 5, mitochondrial serine/threonine protein phosphatase; PHB2: prohibitin 2; PINK1: PTEN induced kinase 1; qPCR: quantitative real-time reverse transcription PCR; Roc-A: Rocaglamide A; PRKN/Parkin: parkin RBR E3 ubiquitin protein ligase; SCVs: Salmonella-containing vacuoles; siRNA: small interfering RNA; SPI-2: Salmonella pathogenicity island 2; SseJ: secretion system effector J; S. Typhimurium: Salmonella enterica serovar Typhimurium; S.T-ΔSseJ: SseJ gene-deleted Salmonella Typhimurium strains; S.T-CΔSseJ: SseJ-complemented Salmonella Typhimurium strains; WT: wild-type.