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Molecular Cell Mar 2021Autophagy deficiency in fed conditions leads to the formation of protein inclusions highlighting the contribution of this lysosomal delivery route to cellular...
Autophagy deficiency in fed conditions leads to the formation of protein inclusions highlighting the contribution of this lysosomal delivery route to cellular proteostasis. Selective autophagy pathways exist that clear accumulated and aggregated ubiquitinated proteins. Receptors for this type of autophagy (aggrephagy) include p62, NBR1, TOLLIP, and OPTN, which possess LC3-interacting regions and ubiquitin-binding domains (UBDs), thus working as a bridge between LC3/GABARAP proteins and ubiquitinated substrates. However, the identity of aggrephagy substrates and the redundancy of aggrephagy and related UBD-containing receptors remains elusive. Here, we combined proximity labeling and organelle enrichment with quantitative proteomics to systematically map the autophagic degradome targeted by UBD-containing receptors under basal and proteostasis-challenging conditions in human cell lines. We identified various autophagy substrates, some of which were differentially engulfed by autophagosomal and endosomal membranes via p62 and TOLLIP, respectively. Overall, this resource will allow dissection of the proteostasis contribution of autophagy to numerous individual proteins.
Topics: Autophagosomes; Autophagy; Autophagy-Related Proteins; HEK293 Cells; HeLa Cells; Humans; Protein Interaction Maps; Proteolysis; Proteomics; Proteostasis; Ubiquitination
PubMed: 33545068
DOI: 10.1016/j.molcel.2021.01.009 -
Proceedings of the National Academy of... Sep 2022Autophagosomes are unique organelles that form de novo as double-membrane vesicles engulfing cytosolic material for destruction. Their biogenesis involves membrane...
Autophagosomes are unique organelles that form de novo as double-membrane vesicles engulfing cytosolic material for destruction. Their biogenesis involves membrane transformations of distinctly shaped intermediates whose ultrastructure is poorly understood. Here, we combine cell biology, correlative cryo-electron tomography (cryo-ET), and extensive data analysis to reveal the step-by-step structural progression of autophagosome biogenesis at high resolution directly within yeast cells. The analysis uncovers an unexpectedly thin intermembrane distance that is dilated at the phagophore rim. Mapping of individual autophagic structures onto a timeline based on geometric features reveals a dynamical change of membrane shape and curvature in growing phagophores. Moreover, our tomograms show the organelle interactome of growing autophagosomes, highlighting a polar organization of contact sites between the phagophore and organelles, such as the vacuole and the endoplasmic reticulum (ER). Collectively, these findings have important implications for the contribution of different membrane sources during autophagy and for the forces shaping and driving phagophores toward closure without a templating cargo.
Topics: Autophagosomes; Cell Membrane; Endoplasmic Reticulum; Macroautophagy; Saccharomyces cerevisiae; Vacuoles
PubMed: 36122245
DOI: 10.1073/pnas.2209823119 -
Autophagy Apr 2024Induction of autophagy is a primordial function of the cGAS-STING pathway. However, the molecular mechanisms regulating autophagosome formation during STING-induced...
Induction of autophagy is a primordial function of the cGAS-STING pathway. However, the molecular mechanisms regulating autophagosome formation during STING-induced autophagy remain largely unknown. Recently, we reported that STING directly interacts with WIPI2 to recruit WIPI2 onto STING-positive vesicles for LC3 lipidation and autophagosome formation. We found that STING and PtdIns3P competitively bind to the FRRG motif of WIPI2, resulting in a mutual inhibition between STING-induced and PtdIns3P-dependent autophagy. We also showed that STING-WIPI2 interaction is necessary for cells to clear cytoplasmic DNA and attenuate activated cGAS-STING signaling. In summary, by identifying the interaction between STING and WIPI2, our study revealed a mechanism that allows STING to bypass the canonical upstream machinery to induce autophagosome formation. ATG: autophagy-related; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; cGAMP: cyclic GMP-AMP; cGAS: cyclic GMP-AMP synthase; ER: endoplasmic reticulum; ERGIC: ER-Golgi intermediate compartment; IRF3: interferon regulatory factor 3; PtdIns3P: phosphatidylinositol-3-phosphate; SQSTM1: sequestosome 1; STING: stimulator of interferon genes; TBK1: TANK-binding kinase 1; ULK1: unc-51 like autophagy activating kinase 1; WIPI2: WD repeat domain, phosphoinositide interacting 2.
Topics: Autophagosomes; Membrane Proteins; Humans; Autophagy; Animals; Signal Transduction; Autophagy-Related Proteins; Phosphate-Binding Proteins
PubMed: 37041719
DOI: 10.1080/15548627.2023.2202108 -
Proceedings of the Japan Academy.... 2020Autophagy is an intracellular degradation system that is present in most eukaryotes. In the process of autophagy, double membrane vesicles called autophagosomes... (Review)
Review
Autophagy is an intracellular degradation system that is present in most eukaryotes. In the process of autophagy, double membrane vesicles called autophagosomes sequester a wide variety of cellular constituents and deliver them to lytic organelles: lysosomes in mammals and vacuoles in yeast and plants. Although autophagy used to be considered a non-selective process in its target sequestration into autophagosomes, recent studies have revealed that autophagosomes can also selectively sequester certain proteins and organelles that have become unnecessary or harmful for the cell. We recently discovered that the endoplasmic reticulum (ER) is degraded by autophagy in a selective manner in the budding yeast Saccharomyces cerevisiae, and identified "receptor proteins" that play pivotal roles in this "ER-phagy" pathway. Moreover, several ER-phagy receptors in mammalian cells have also been reported. This report provides an overview of our current knowledge on ER-phagy and discuss their mechanisms, physiological roles, and possible links to human diseases.
Topics: Animals; Autophagosomes; Autophagy; Endoplasmic Reticulum; Endoplasmic Reticulum Stress; Humans; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins
PubMed: 31932525
DOI: 10.2183/pjab.96.001 -
Autophagy Jan 2022Ion exchange between intracellular and extracellular spaces is the basic mechanism for controlling cell metabolism and signal transduction. This process is mediated by...
Ion exchange between intracellular and extracellular spaces is the basic mechanism for controlling cell metabolism and signal transduction. This process is mediated by ion channels and transporters on the plasma membrane, or intracellular membranes that surround various organelles, in response to environmental stimuli. Macroautophagy (hereafter referred to as autophagy) is one of the lysosomal-dependent degradation pathways that maintains homeostasis through the degradation and recycling of cellular components (e.g., dysfunctional proteins and damaged organelles). Although autophagy-related (ATG) proteins play a central role in regulating the formation of autophagy-related member structures (e.g., phagophores, autophagosomes, and autolysosomes), the autophagic process also involves changes in expression and function of ion channels and transporters. Here we discuss current knowledge of the mechanisms that regulate autophagy in mammalian cells, with special attention to the ion channels and transporters. We also highlight prospects for the development of drugs targeting ion channels and transporters in autophagy.
Topics: Animals; Autophagosomes; Autophagy; Intracellular Membranes; Ion Channels; Lysosomes; Mammals
PubMed: 33657975
DOI: 10.1080/15548627.2021.1885147 -
Biochemical Society Transactions Feb 2022Autophagy is an evolutionally conserved cytoplasmic degradation pathway in which the double membrane structure, autophagosome sequesters cytoplasmic material and... (Review)
Review
Autophagy is an evolutionally conserved cytoplasmic degradation pathway in which the double membrane structure, autophagosome sequesters cytoplasmic material and delivers them to lysosomes for degradation. Many autophagy related (ATG) proteins participate in the regulation of the several steps of autophagic process. Among ATGs, ubiquitin-like protein, ATG8 plays a pivotal role in autophagy. ATG8 is directly conjugated on lipid in autophagosome membrane upon induction of autophagy thus providing a good marker to monitor and analyze autophagy process. However, recent discoveries suggest that ATG8 has autophagy independent non-canonical functions and ATG8 positive structures are not always autophagosomes. This review briefly overviews canonical and non-canonical roles of ATG8 and introduce novel function of ATG8 to activate Transcriptional Factor EB(TFEB), a master transcription factor of autophagy and lysosome function during lysosomal damage.
Topics: Autophagosomes; Autophagy; Autophagy-Related Protein 8 Family; Lysosomes; Transcription Factors
PubMed: 35166325
DOI: 10.1042/BST20210813 -
Cells Mar 2020Autophagy is a catabolic process involving vacuolar sequestration of intracellular components and their targeting to lysosomes for degradation, thus supporting nutrient... (Review)
Review
Autophagy is a catabolic process involving vacuolar sequestration of intracellular components and their targeting to lysosomes for degradation, thus supporting nutrient recycling and energy regeneration. Accumulating evidence indicates that in addition to being a bulk, nonselective degradation mechanism, autophagy may selectively eliminate damaged mitochondria to promote mitochondrial turnover, a process termed "mitophagy". Mitophagy sequesters dysfunctional mitochondria via ubiquitination and cargo receptor recognition and has emerged as an important event in the regulation of liver physiology. Recent studies have shown that mitophagy may participate in the pathogenesis of various liver diseases, such as liver injury, liver steatosis/fatty liver disease, hepatocellular carcinoma, viral hepatitis, and hepatic fibrosis. This review summarizes the current knowledge on the molecular regulations and functions of mitophagy in liver physiology and the roles of mitophagy in the development of liver-related diseases. Furthermore, the therapeutic implications of targeting hepatic mitophagy to design a new strategy to cure liver diseases are discussed.
Topics: Autophagosomes; Autophagy; Humans; Liver; Liver Diseases; Mitophagy; Translational Research, Biomedical
PubMed: 32235615
DOI: 10.3390/cells9040831 -
Nature Reviews. Molecular Cell Biology Mar 2021
Topics: Autophagosomes; Autophagy; Cytosol; Phagosomes
PubMed: 33495650
DOI: 10.1038/s41580-020-00321-x -
Journal of Molecular and Cellular... Apr 2022Autophagy mediates cellular quality control mechanisms and energy homeostasis through lysosomal degradation. Autophagy is typically viewed as an adaptive process that... (Review)
Review
Autophagy mediates cellular quality control mechanisms and energy homeostasis through lysosomal degradation. Autophagy is typically viewed as an adaptive process that allows cells to survive against stress, such as nutrient deprivation and hypoxia. However, autophagy also mediates cell death during development and in response to stress. Cell death accompanied by autophagy activation and accumulation of autophagosomes has been classified as type II programmed cell death. Compared to the wealth of knowledge regarding the adaptive role of autophagy, however, the molecular mechanisms through which autophagy induces cell death and its functional significance are poorly understood. Autophagy is activated excessively under some conditions, causing uncontrolled degradation of cellular materials and cell death. An imbalance between autophagosome formation and lysosomal degradation causes a massive accumulation of autophagosomes, which subsequently causes cellular dysfunction and death. Dysregulation of autophagy induces a unique form of cell death, termed autosis, with defined morphological and biochemical features distinct from other forms of programmed cell death, such as apoptosis and necrosis. In the heart, dysregulated autophagy induces death of cardiomyocytes and actively mediates cardiac injury and dysfunction in some conditions, including reperfusion injury, doxorubicin cardiomyopathy, and lysosomal storage disorders. The goal in this review is to introduce the concept of autophagic cell death and discuss its functional significance in various cardiac conditions.
Topics: Apoptosis; Autophagosomes; Autophagy; Lysosomes; Myocytes, Cardiac
PubMed: 34919896
DOI: 10.1016/j.yjmcc.2021.12.006 -
Nature Communications Nov 2023Autophagosomes are double-membrane vesicles generated intracellularly to encapsulate substrates for lysosomal degradation during autophagy. Phase separated p62 body...
Autophagosomes are double-membrane vesicles generated intracellularly to encapsulate substrates for lysosomal degradation during autophagy. Phase separated p62 body plays pivotal roles during autophagosome formation, however, the underlying mechanisms are still not fully understood. Here we describe a spatial membrane gathering mode by which p62 body functions in autophagosome formation. Mass spectrometry-based proteomics reveals significant enrichment of vesicle trafficking components within p62 body. Combining cellular experiments and biochemical reconstitution assays, we confirm the gathering of ATG9 and ATG16L1-positive vesicles around p62 body, especially in Atg2ab DKO cells with blocked lipid transfer and vesicle fusion. Interestingly, p62 body also regulates ATG9 and ATG16L vesicle trafficking flux intracellularly. We further determine the lipid contents associated with p62 body via lipidomic profiling. Moreover, with in vitro kinase assay, we uncover the functions of p62 body as a platform to assemble ULK1 complex and invigorate PI3KC3-C1 kinase cascade for PI3P generation. Collectively, our study raises a membrane-based working model for multifaceted p62 body in controlling autophagosome biogenesis, and highlights the interplay between membraneless condensates and membrane vesicles in regulating cellular functions.
Topics: Autophagosomes; Autophagy; Macroautophagy; Phagosomes; Autophagy-Related Proteins; Lipids
PubMed: 37957156
DOI: 10.1038/s41467-023-42829-8