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Proceedings of the National Academy of... Aug 2023Cells maintain optimal levels of lysosome degradative activity to protect against pathogens, clear waste, and generate nutrients. Here, we show that LRRK2, a protein...
Cells maintain optimal levels of lysosome degradative activity to protect against pathogens, clear waste, and generate nutrients. Here, we show that LRRK2, a protein that is tightly linked to Parkinson's disease, negatively regulates lysosome degradative activity in macrophages and microglia via a transcriptional mechanism. Depletion of LRRK2 and inhibition of LRRK2 kinase activity enhanced lysosomal proteolytic activity and increased the expression of multiple lysosomal hydrolases. Conversely, the kinase hyperactive LRRK2 G2019S Parkinson's disease mutant suppressed lysosomal degradative activity and gene expression. We identified MiT-TFE transcription factors (TFE3, TFEB, and MITF) as mediators of LRRK2-dependent control of lysosomal gene expression. LRRK2 negatively regulated the abundance and nuclear localization of these transcription factors and their depletion prevented LRRK2-dependent changes in lysosome protein levels. These observations define a role for LRRK2 in controlling lysosome degradative activity and support a model wherein LRRK2 hyperactivity may increase Parkinson's disease risk by suppressing lysosome degradative activity.
Topics: Humans; Transcription Factors; Parkinson Disease; Microglia; Leucine-Rich Repeat Serine-Threonine Protein Kinase-2; Lysosomes; Macrophages
PubMed: 37487100
DOI: 10.1073/pnas.2303789120 -
Nature Communications Jul 2023Differentiation is critical for cell fate decisions, but the signals involved remain unclear. The kidney proximal tubule (PT) cells reabsorb disulphide-rich proteins...
Differentiation is critical for cell fate decisions, but the signals involved remain unclear. The kidney proximal tubule (PT) cells reabsorb disulphide-rich proteins through endocytosis, generating cystine via lysosomal proteolysis. Here we report that defective cystine mobilization from lysosomes through cystinosin (CTNS), which is mutated in cystinosis, diverts PT cells towards growth and proliferation, disrupting their functions. Mechanistically, cystine storage stimulates Ragulator-Rag GTPase-dependent recruitment of mechanistic target of rapamycin complex 1 (mTORC1) and its constitutive activation. Re-introduction of CTNS restores nutrient-dependent regulation of mTORC1 in knockout cells, whereas cell-permeant analogues of L-cystine, accumulating within lysosomes, render wild-type cells resistant to nutrient withdrawal. Therapeutic mTORC1 inhibition corrects lysosome and differentiation downstream of cystine storage, and phenotypes in preclinical models of cystinosis. Thus, cystine serves as a lysosomal signal that tailors mTORC1 and metabolism to direct epithelial cell fate decisions. These results identify mechanisms and therapeutic targets for dysregulated homeostasis in cystinosis.
Topics: Humans; Cystine; Cystinosis; Mechanistic Target of Rapamycin Complex 1; Kidney; Epithelial Cells; Lysosomes; Amino Acid Transport Systems, Neutral
PubMed: 37452023
DOI: 10.1038/s41467-023-39261-3 -
The Journal of Experimental Medicine Nov 2023Accumulation of lipotoxic lipids, such as free cholesterol, induces hepatocyte death and subsequent inflammation and fibrosis in the pathogenesis of nonalcoholic...
Accumulation of lipotoxic lipids, such as free cholesterol, induces hepatocyte death and subsequent inflammation and fibrosis in the pathogenesis of nonalcoholic steatohepatitis (NASH). However, the underlying mechanisms remain unclear. We have previously reported that hepatocyte death locally induces phenotypic changes in the macrophages surrounding the corpse and remnant lipids, thereby promoting liver fibrosis in a murine model of NASH. Here, we demonstrated that lysosomal cholesterol overload triggers lysosomal dysfunction and profibrotic activation of macrophages during the development of NASH. β-cyclodextrin polyrotaxane (βCD-PRX), a unique supramolecule, is designed to elicit free cholesterol from lysosomes. Treatment with βCD-PRX ameliorated cholesterol accumulation and profibrotic activation of macrophages surrounding dead hepatocytes with cholesterol crystals, thereby suppressing liver fibrosis in a NASH model, without affecting the hepatic cholesterol levels. In vitro experiments revealed that cholesterol-induced lysosomal stress triggered profibrotic activation in macrophages predisposed to the steatotic microenvironment. This study provides evidence that dysregulated cholesterol metabolism in macrophages would be a novel mechanism of NASH.
Topics: Animals; Mice; Non-alcoholic Fatty Liver Disease; Disease Models, Animal; Liver Cirrhosis; Macrophages; Cholesterol; Lysosomes
PubMed: 37725372
DOI: 10.1084/jem.20220681 -
Nature Communications Oct 2023Toll-like receptors (TLRs) are a class of proteins that play critical roles in recognizing pathogens and initiating innate immune responses. TASL, a recently identified...
Toll-like receptors (TLRs) are a class of proteins that play critical roles in recognizing pathogens and initiating innate immune responses. TASL, a recently identified innate immune adaptor protein for endolysosomal TLR7/8/9 signaling, is recruited by the lysosomal proton-coupled amino-acid transporter SLC15A4, and then activates IRF5, which in turn triggers the transcription of type I interferons and cytokines. Here, we report three cryo-electron microscopy (cryo-EM) structures of human SLC15A4 in the apo monomeric and dimeric state and as a TASL-bound complex. The apo forms are in an outward-facing conformation, with the dimeric form showing an extensive interface involving four cholesterol molecules. The structure of the TASL-bound complex reveals an unprecedented interaction mode with solute carriers. During the recruitment of TASL, SLC15A4 undergoes a conformational change from an outward-facing, lysosomal lumen-exposed state to an inward-facing state to form a binding pocket, allowing the N-terminal helix of TASL to be inserted into. Our findings provide insights into the molecular basis of regulatory switch involving a human solute carrier and offers an important framework for structure-guided drug discovery targeting SLC15A4-TASL-related human autoimmune diseases.
Topics: Humans; Cryoelectron Microscopy; Signal Transduction; Toll-Like Receptors; Immunity, Innate; Lysosomes; Nerve Tissue Proteins; Membrane Transport Proteins
PubMed: 37863913
DOI: 10.1038/s41467-023-42210-9 -
Cancer Communications (London, England) Jan 2024Transmembrane 4 L six family member 5 (TM4SF5) translocates subcellularly and functions metabolically, although it is unclear how intracellular TM4SF5 translocation is...
BACKGROUND
Transmembrane 4 L six family member 5 (TM4SF5) translocates subcellularly and functions metabolically, although it is unclear how intracellular TM4SF5 translocation is linked to metabolic contexts. It is thus of interests to understand how the traffic dynamics of TM4SF5 to subcellular endosomal membranes are correlated to regulatory roles of metabolisms.
METHODS
Here, we explored the metabolic significance of TM4SF5 localization at mitochondria-lysosome contact sites (MLCSs), using in vitro cells and in vivo animal systems, via approaches by immunofluorescence, proximity labelling based proteomics analysis, organelle reconstitution etc. RESULTS: Upon extracellular glucose repletion following depletion, TM4SF5 became enriched at MLCSs via an interaction between mitochondrial FK506-binding protein 8 (FKBP8) and lysosomal TM4SF5. Proximity labeling showed molecular clustering of phospho-dynamic-related protein I (DRP1) and certain mitophagy receptors at TM4SF5-enriched MLCSs, leading to mitochondrial fission and autophagy. TM4SF5 bound NPC intracellular cholesterol transporter 1 (NPC1) and free cholesterol, and mediated export of lysosomal cholesterol to mitochondria, leading to impaired oxidative phosphorylation but intact tricarboxylic acid (TCA) cycle and β-oxidation. In mouse models, hepatocyte Tm4sf5 promoted mitophagy and cholesterol transport to mitochondria, both with positive relations to liver malignancy.
CONCLUSIONS
Our findings suggested that TM4SF5-enriched MLCSs regulate glucose catabolism by facilitating cholesterol export for mitochondrial reprogramming, presumably while hepatocellular carcinogenesis, recapitulating aspects for hepatocellular carcinoma metabolism with mitochondrial reprogramming to support biomolecule synthesis in addition to glycolytic energetics.
Topics: Animals; Mice; Membrane Proteins; Cell Movement; Mitochondria; Lysosomes; Cholesterol
PubMed: 38133457
DOI: 10.1002/cac2.12510 -
Cell Calcium Jul 2023Multiple forms of regulated cell death (RCD) have been characterized, each of which originates from the activation of a dedicated molecular machinery. RCD can occur in... (Review)
Review
Multiple forms of regulated cell death (RCD) have been characterized, each of which originates from the activation of a dedicated molecular machinery. RCD can occur in purely physiological settings or upon failing cellular adaptation to stress. Caions have been shown to physically interact with - and hence regulate - various components of the RCD machinery. Moreover, intracellular Ca accumulation can promote organellar dysfunction to degree that can be overtly cytotoxic or sensitize cells to RCD elicited by other stressors. Here, we provide an overview of the main links between Caand different forms of RCD, including apoptosis, mitochondrial permeability transition (MPT)-driven necrosis, necroptosis, ferroptosis, lysosome-dependent cell death, and parthanatos.
Topics: Humans; Apoptosis; Cell Death; Lysosomes; Necrosis; Signal Transduction
PubMed: 37210868
DOI: 10.1016/j.ceca.2023.102759 -
Nature Cancer May 2024Pancreatic ductal adenocarcinoma is a highly metastatic disease and macrophages support liver metastases. Efferocytosis, or engulfment of apoptotic cells by macrophages,...
Pancreatic ductal adenocarcinoma is a highly metastatic disease and macrophages support liver metastases. Efferocytosis, or engulfment of apoptotic cells by macrophages, is an essential process in tissue homeostasis and wound healing, but its role in metastasis is less well understood. Here, we found that the colonization of the hepatic metastatic site is accompanied by low-grade tissue injury and that efferocytosis-mediated clearance of parenchymal dead cells promotes macrophage reprogramming and liver metastasis. Mechanistically, progranulin expression in macrophages is necessary for efficient efferocytosis by controlling lysosomal acidification via cystic fibrosis transmembrane conductance regulator and the degradation of lysosomal cargo, resulting in LXRα/RXRα-mediated macrophage conversion and upregulation of arginase 1. Pharmacological blockade of efferocytosis or macrophage-specific genetic depletion of progranulin impairs macrophage conversion, improves CD8 T cell functions, and reduces liver metastasis. Our findings reveal how hard-wired functions of macrophages in tissue repair contribute to liver metastasis and identify potential targets for prevention of pancreatic ductal adenocarcinoma liver metastasis.
Topics: Tumor Microenvironment; Pancreatic Neoplasms; Humans; Liver Neoplasms; Animals; Mice; Macrophages; Carcinoma, Pancreatic Ductal; Phagocytosis; Cell Line, Tumor; CD8-Positive T-Lymphocytes; Apoptosis; Lysosomes; Arginase; Efferocytosis
PubMed: 38355776
DOI: 10.1038/s43018-024-00731-2 -
Science Advances Jul 2023Lysosome dysfunction arises early and propels Alzheimer's disease (AD). Herein, we show that amyloid precursor protein (APP), linked to early-onset AD in Down syndrome...
Lysosome dysfunction arises early and propels Alzheimer's disease (AD). Herein, we show that amyloid precursor protein (APP), linked to early-onset AD in Down syndrome (DS), acts directly via its β-C-terminal fragment (βCTF) to disrupt lysosomal vacuolar (H)-adenosine triphosphatase (v-ATPase) and acidification. In human DS fibroblasts, the phosphorylated YENPTY internalization motif of APP-βCTF binds selectively within a pocket of the v-ATPase V0a1 subunit cytoplasmic domain and competitively inhibits association of the V1 subcomplex of v-ATPase, thereby reducing its activity. Lowering APP-βCTF Tyr phosphorylation restores v-ATPase and lysosome function in DS fibroblasts and in vivo in brains of DS model mice. Notably, lowering APP-βCTF Tyr phosphorylation below normal constitutive levels boosts v-ATPase assembly and activity, suggesting that v-ATPase may also be modulated tonically by phospho-APP-βCTF. Elevated APP-βCTF Tyr phosphorylation in two mouse AD models similarly disrupts v-ATPase function. These findings offer previously unknown insight into the pathogenic mechanism underlying faulty lysosomes in all forms of AD.
Topics: Mice; Humans; Animals; Amyloid beta-Protein Precursor; Down Syndrome; Alzheimer Disease; Adenosine Triphosphatases; Lysosomes; Disease Models, Animal; Amyloid beta-Peptides
PubMed: 37494443
DOI: 10.1126/sciadv.adg1925 -
Nature Communications Jul 2023Batten disease, one of the most devastating types of neurodegenerative lysosomal storage disorders, is caused by mutations in CLN3. Here, we show that CLN3 is a...
Batten disease, one of the most devastating types of neurodegenerative lysosomal storage disorders, is caused by mutations in CLN3. Here, we show that CLN3 is a vesicular trafficking hub connecting the Golgi and lysosome compartments. Proteomic analysis reveals that CLN3 interacts with several endo-lysosomal trafficking proteins, including the cation-independent mannose 6 phosphate receptor (CI-M6PR), which coordinates the targeting of lysosomal enzymes to lysosomes. CLN3 depletion results in mis-trafficking of CI-M6PR, mis-sorting of lysosomal enzymes, and defective autophagic lysosomal reformation. Conversely, CLN3 overexpression promotes the formation of multiple lysosomal tubules, which are autophagy and CI-M6PR-dependent, generating newly formed proto-lysosomes. Together, our findings reveal that CLN3 functions as a link between the M6P-dependent trafficking of lysosomal enzymes and lysosomal reformation pathway, explaining the global impairment of lysosomal function in Batten disease.
Topics: Humans; Membrane Glycoproteins; Neuronal Ceroid-Lipofuscinoses; Receptor, IGF Type 2; Proteomics; Molecular Chaperones; Lysosomes; Hydrolases; Autophagy
PubMed: 37400440
DOI: 10.1038/s41467-023-39643-7 -
The Journal of Clinical Investigation Aug 2023Protein aggregation is a hallmark of many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS). Although mutations in TARDBP, encoding transactive...
Protein aggregation is a hallmark of many neurodegenerative disorders, including amyotrophic lateral sclerosis (ALS). Although mutations in TARDBP, encoding transactive response DNA-binding protein 43 kDa (TDP-43), account for less than 1% of all ALS cases, TDP-43-positive aggregates are present in nearly all ALS patients, including patients with sporadic ALS (sALS) or carrying other familial ALS-causing (fALS-causing) mutations. Interestingly, TDP-43 inclusions are also present in subsets of patients with frontotemporal dementia, Alzheimer's disease, and Parkinson's disease; therefore, methods of activating intracellular protein quality control machinery capable of clearing toxic cytoplasmic TDP-43 species may alleviate disease-related phenotypes. Here, we identify a function of nemo-like kinase (Nlk) as a negative regulator of lysosome biogenesis. Genetic or pharmacological reduction of Nlk increased lysosome formation and improved clearance of aggregated TDP-43. Furthermore, Nlk reduction ameliorated pathological, behavioral, and life span deficits in 2 distinct mouse models of TDP-43 proteinopathy. Because many toxic proteins can be cleared through the autophagy/lysosome pathway, targeted reduction of Nlk represents a potential approach to therapy development for multiple neurodegenerative disorders.
Topics: Animals; Mice; Amyotrophic Lateral Sclerosis; DNA-Binding Proteins; Lysosomes; Neurodegenerative Diseases; Humans
PubMed: 37384409
DOI: 10.1172/JCI138207