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Traffic (Copenhagen, Denmark) May 2022Since the discovery of lysosomes more than 70 years ago, much has been learned about the functions of these organelles. Lysosomes were regarded as exclusively... (Review)
Review
Since the discovery of lysosomes more than 70 years ago, much has been learned about the functions of these organelles. Lysosomes were regarded as exclusively degradative organelles, but more recent research has shown that they play essential roles in several other cellular functions, such as nutrient sensing, intracellular signalling and metabolism. Methodological advances played a key part in generating our current knowledge about the biology of this multifaceted organelle. In this review, we cover current methods used to analyze lysosome morphology, positioning, motility and function. We highlight the principles behind these methods, the methodological strategies and their advantages and limitations. To extract accurate information and avoid misinterpretations, we discuss the best strategies to identify lysosomes and assess their characteristics and functions. With this review, we aim to stimulate an increase in the quantity and quality of research on lysosomes and further ground-breaking discoveries on an organelle that continues to surprise and excite cell biologists.
Topics: Lysosomes; Metabolic Networks and Pathways; Signal Transduction
PubMed: 35343629
DOI: 10.1111/tra.12839 -
Nature Cell Biology Apr 2022Autolysosomes contain components from autophagosomes and lysosomes. The contents inside the autolysosomal lumen are degraded during autophagy, while the fate of...
Autolysosomes contain components from autophagosomes and lysosomes. The contents inside the autolysosomal lumen are degraded during autophagy, while the fate of autophagosomal components on the autolysosomal membrane remains unknown. Here we report that the autophagosomal membrane components are not degraded, but recycled from autolysosomes through a process coined in this study as autophagosomal components recycling (ACR). We further identified a multiprotein complex composed of SNX4, SNX5 and SNX17 essential for ACR, which we termed 'recycler'. In this, SNX4 and SNX5 form a heterodimer that recognizes autophagosomal membrane proteins and is required for generating membrane curvature on autolysosomes, both via their BAR domains, to mediate the cargo sorting process. SNX17 interacts with both the dynein-dynactin complex and the SNX4-SNX5 dimer to facilitate the retrieval of autophagosomal membrane components. Our discovery of ACR and identification of the recycler reveal an important retrieval and recycling pathway on autolysosomes.
Topics: Autophagosomes; Autophagy; Dyneins; Lysosomes; Protein Transport
PubMed: 35332264
DOI: 10.1038/s41556-022-00861-8 -
The Journal of Cell Biology Jun 2022The endolysosome system plays central roles in both autophagic degradation and secretory pathways, including the release of extracellular vesicles and particles (EVPs)....
The endolysosome system plays central roles in both autophagic degradation and secretory pathways, including the release of extracellular vesicles and particles (EVPs). Although previous work reveals important interconnections between autophagy and EVP-mediated secretion, our understanding of these secretory events during endolysosome inhibition remains incomplete. Here, we delineate a secretory autophagy pathway upregulated in response to endolysosomal inhibition, which mediates EVP-associated release of autophagic cargo receptors, including p62/SQSTM1. This secretion is highly regulated and dependent on multiple ATGs required for autophagosome formation, as well as the small GTPase Rab27a. Furthermore, disrupting autophagosome maturation, either via genetic inhibition of autophagosome-to-autolysosome fusion or expression of SARS-CoV-2 ORF3a, is sufficient to induce EVP secretion of autophagy cargo receptors. Finally, ATG-dependent EVP secretion buffers against the intracellular accumulation of autophagy cargo receptors when classical autophagic degradation is impaired. Thus, we propose secretory autophagy via EVPs functions as an alternate route to clear sequestered material and maintain proteostasis during endolysosomal dysfunction or impaired autophagosome maturation.
Topics: Autophagosomes; Autophagy; Extracellular Vesicles; Humans; Lysosomes; Proteostasis; SARS-CoV-2; Sequestosome-1 Protein; Viroporin Proteins; rab27 GTP-Binding Proteins
PubMed: 35446347
DOI: 10.1083/jcb.202110151 -
Molecular Genetics and Metabolism Feb 2021
Topics: Humans; Lysosomes; Metabolic Diseases
PubMed: 33541581
DOI: 10.1016/j.ymgme.2020.12.293 -
Angewandte Chemie (International Ed. in... May 2021The ability to regulate membrane protein abundance offers great opportunities for developing therapeutic sites for various diseases. Herein, we describe a platform for...
The ability to regulate membrane protein abundance offers great opportunities for developing therapeutic sites for various diseases. Herein, we describe a platform for the targeted degradation of membrane-associated proteins using bispecific aptamer chimeras that bind both the cell-surface lysosome-shuttling receptor (IGFIIR) and the targeted membrane-bound proteins of interest. We demonstrate that the aptamer chimeras can efficiently and quickly shuttle the therapeutically relevant membrane proteins of Met and PTK-7 to lysosomes and degrade them through the lysosomal protein degradation machinery. We anticipate that our method will provide a universal platform for the use of readily synthesized aptamer materials for biochemical research and potential therapeutics.
Topics: Aptamers, Nucleotide; Cell Membrane; HeLa Cells; Humans; Lysosomes; Membrane Proteins
PubMed: 33634555
DOI: 10.1002/anie.202102170 -
Autophagy Oct 2019Multiple sources contribute membrane and protein machineries to construct functional macroautophagic/autophagic structures. However, the underlying molecular mechanisms...
Multiple sources contribute membrane and protein machineries to construct functional macroautophagic/autophagic structures. However, the underlying molecular mechanisms remain elusive. Here, we show that RAB2 connects the Golgi network to autophagy pathway by delivering membrane and by sequentially engaging distinct autophagy machineries. In unstressed cells, RAB2 resides primarily in the Golgi apparatus, as evidenced by its interaction and colocalization with GOLGA2/GM130. Importantly, autophagy stimuli dissociate RAB2 from GOLGA2 to interact with ULK1 complex, which facilitates the recruitment of ULK1 complex to form phagophores. Intriguingly, RAB2 appears to modulate ULK1 kinase activity to propagate signals for autophagosome formation. Subsequently, RAB2 switches to interact with autophagosomal RUBCNL/PACER and STX17 to further specify the recruitment of HOPS complex for autolysosome formation. Together, our study reveals a multivalent pathway in bulk autophagy regulation, and provides mechanistic insights into how the Golgi apparatus contributes to the formation of different autophagic structures. ACTB: actin beta; ATG9: autophagy related 9A; ATG14: autophagy related 14; ATG16L1: autophagy related 16 like 1; BCAP31: B cell receptor associated protein 31; BECN1: beclin 1; Ctrl: control; CQ: chloroquine; CTSD: cathepsin D; DMSO: dimethyl sulfoxide; EBSS: Earle's balanced salt solution; EEA1: early endosome antigen 1; GDI: guanine nucleotide dissociation inhibitor; GFP: green fluorescent protein; GOLGA2: golgin A2; HOPS: homotypic fusion and protein sorting complex; IP: immunoprecipitation; KD: knockdown; KO: knockout; LAMP1: lysosomal associated membrane protein 1; LC3: microtubule-associated protein 1 light chain 3; OE: overexpression; PtdIns3K: class III phosphatidylinositol 3-kinase; SQSTM1/p62: sequestosome 1; RAB2: RAB2A, member RAS oncogene family; RAB7: RAB7A, member RAS oncogene family; RAB11: RAB11A, member RAS oncogene family; RUBCNL/PACER: rubicon like autophagy enhancer; STX17: syntaxin 17; TBC1D14: TBC1 domain family member 14; TFRC: transferrin receptor; TGOLN2: trans-golgi network protein 2; TUBB: tubulin beta class I; ULK1: unc-51 like autophagy activating kinase 1; VPS41: VPS41, HOPS complex subunit; WB: western blot; WT: wild type; YPT1: GTP-binding protein YPT1.
Topics: Animals; Autophagosomes; Autophagy; Cells, Cultured; Eukaryotic Cells; HEK293 Cells; HeLa Cells; Humans; Lysosomes; Male; Mammals; Mice; Mice, Inbred C57BL; Mice, Knockout; rab2 GTP-Binding Protein
PubMed: 30957628
DOI: 10.1080/15548627.2019.1596478 -
Autophagy Oct 2023Inter-organelle contacts enable crosstalk among organelles, facilitating the exchange of materials and coordination of cellular events. In this study, we demonstrated...
Inter-organelle contacts enable crosstalk among organelles, facilitating the exchange of materials and coordination of cellular events. In this study, we demonstrated that, upon starvation, autolysosomes recruit Pi4KIIα (Phosphatidylinositol 4-kinase II α) to generate phosphatidylinositol-4-phosphate (PtdIns4P) on their surface and establish endoplasmic reticulum (ER)-autolysosome contacts through PtdIns4P binding proteins Osbp (Oxysterol binding protein) and cert (ceramide transfer protein). We found that the Sac1 (Sac1 phosphatase), Osbp, and cert proteins are required for the reduction of PtdIns4P on autolysosomes. Loss of any of these proteins leads to defective macroautophagy/autophagy and neurodegeneration. Osbp, cert, and Sac1 are required for ER-Golgi contacts in fed cells. Our data establishes a new mode of organelle contact formation - the ER-Golgi contact machinery can be reused by ER-autolysosome contacts by re-locating PtdIns4P from the Golgi apparatus to autolysosomes when faced with starvation. Atg1: Autophagy-related 1; Atg8: Autophagy-related 8; Atg9: Autophagy-related 9; Atg12: Autophagy-related 12; cert: ceramide transfer protein; Cp1/CathL: cysteine proteinase-1; CTL: control; ER: endoplasmic reticulum; ERMCS: ER-mitochondria contact site; fwd: four wheel drive; GM130: Golgi matrix protein 130 kD; Osbp: Oxysterol binding protein; PG: phagophore; PtdIns4K: phosphatidylinositol 4-kinase; Pi4KIIα: Phosphatidylinositol 4-kinase II α; Pi4KIIIα: Phosphatidylinositol 4-kinase III α; PtdIns4P: phosphatidylinositol-4-phosphate; PR: photoreceptor cell; RT: room temperature; Sac1: Sac1 phosphatase; Stv: starvation; Syx17: Syntaxin 17; TEM: transmission electron microscopy; VAP: VAMP-associated protein.
Topics: 1-Phosphatidylinositol 4-Kinase; Autophagy; Endoplasmic Reticulum; Lysosomes; Carrier Proteins; Homeostasis; Ceramides; Phosphoric Monoester Hydrolases
PubMed: 37289040
DOI: 10.1080/15548627.2023.2222556 -
Circulation Apr 2016The clinical use of doxorubicin is limited by cardiotoxicity. Histopathological changes include interstitial myocardial fibrosis and the appearance of vacuolated...
BACKGROUND
The clinical use of doxorubicin is limited by cardiotoxicity. Histopathological changes include interstitial myocardial fibrosis and the appearance of vacuolated cardiomyocytes. Whereas dysregulation of autophagy in the myocardium has been implicated in a variety of cardiovascular diseases, the role of autophagy in doxorubicin cardiomyopathy remains poorly defined.
METHODS AND RESULTS
Most models of doxorubicin cardiotoxicity involve intraperitoneal injection of high-dose drug, which elicits lethargy, anorexia, weight loss, and peritoneal fibrosis, all of which confound the interpretation of autophagy. Given this, we first established a model that provokes modest and progressive cardiotoxicity without constitutional symptoms, reminiscent of the effects seen in patients. We report that doxorubicin blocks cardiomyocyte autophagic flux in vivo and in cardiomyocytes in culture. This block was accompanied by robust accumulation of undegraded autolysosomes. We go on to localize the site of block as a defect in lysosome acidification. To test the functional relevance of doxorubicin-triggered autolysosome accumulation, we studied animals with diminished autophagic activity resulting from haploinsufficiency for Beclin 1. Beclin 1(+/-) mice exposed to doxorubicin were protected in terms of structural and functional changes within the myocardium. Conversely, animals overexpressing Beclin 1 manifested an amplified cardiotoxic response.
CONCLUSIONS
Doxorubicin blocks autophagic flux in cardiomyocytes by impairing lysosome acidification and lysosomal function. Reducing autophagy initiation protects against doxorubicin cardiotoxicity.
Topics: Animals; Antibiotics, Antineoplastic; Autophagy; Cell Line; Cells, Cultured; Doxorubicin; Humans; Hydrogen-Ion Concentration; Lysosomes; Mice; Mice, Inbred C57BL; Myocytes, Cardiac; Rats; Rats, Sprague-Dawley
PubMed: 26984939
DOI: 10.1161/CIRCULATIONAHA.115.017443 -
Biochimica Et Biophysica Acta.... Mar 2020Autophagy (greek auto: self; phagein: eating) is a highly conserved process within eukaryotes that degrades long-lived proteins and organelles within lysosomes. Its... (Review)
Review
Autophagy (greek auto: self; phagein: eating) is a highly conserved process within eukaryotes that degrades long-lived proteins and organelles within lysosomes. Its accurate and constant operation in basal conditions ensures cellular homeostasis by degrading damaged cellular components and thereby acting not only as a quality control but as well as an energy supplier. An increasing body of evidence indicates a major role of autophagy in the regulation of cardiac homeostasis and function. In this review, we describe the different forms of mammalian autophagy, their regulations and monitoring with a specific emphasis on the heart. Furthermore, we address the role of autophagy in several forms of cardiomyopathy and the options for therapy.
Topics: Autophagy; Cardiomyopathies; Energy Metabolism; Homeostasis; Humans; Lysosomes; Proteolysis
PubMed: 30831130
DOI: 10.1016/j.bbamcr.2019.01.013 -
Traffic (Copenhagen, Denmark) Jan 2020Lysosomes are key cellular catabolic centers that also perform fundamental metabolic, signaling and quality control functions. Lysosomes are not static and they respond... (Review)
Review
Lysosomes are key cellular catabolic centers that also perform fundamental metabolic, signaling and quality control functions. Lysosomes are not static and they respond dynamically to intra- and extracellular stimuli triggering changes in organelle numbers, size and position. Such physical changes have a strong impact on lysosomal activity ultimately influencing cellular homeostasis. In this review, we summarize the current knowledge on lysosomal size regulation, on its physiological role(s) and association to several disease conditions.
Topics: Autophagy; Homeostasis; Lysosomes; Signal Transduction
PubMed: 31808235
DOI: 10.1111/tra.12714