ABSTRACT
Every cell must satisfy basic requirements for nutrient sensing, utilization and recycling through macromolecular breakdown to coordinate programmes for growth, repair and stress adaptation. The lysosome orchestrates these key functions through the synchronised interplay between hydrolytic enzymes, nutrient transporters and signalling factors, which together enable metabolic coordination with other organelles and regulation of specific gene expression programmes. In this Review, we discuss recent findings on lysosome-dependent signalling pathways, focusing on how the lysosome senses nutrient availability through its physical and functional association with mechanistic target of rapamycin complex 1 (mTORC1) and how, in response, the microphthalmia/transcription factor E (MiT/TFE) transcription factors exert feedback regulation on lysosome biogenesis. We also highlight the emerging interactions of lysosomes with other organelles, which contribute to cellular homeostasis. Lastly, we discuss how lysosome dysfunction contributes to diverse disease pathologies and how inherited mutations that compromise lysosomal hydrolysis, transport or signalling components lead to multi-organ disorders with severe metabolic and neurological impact. A deeper comprehension of lysosomal composition and function, at both the cellular and organismal level, may uncover fundamental insights into human physiology and disease.
Subject(s)
Lysosomes , Signal Transduction , Humans , Signal Transduction/physiology , Mechanistic Target of Rapamycin Complex 1/metabolism , Lysosomes/metabolism , Homeostasis/physiology , Autophagy/physiologyABSTRACT
The lysosome has long been viewed as the recycling center of the cell. However, recent discoveries have challenged this simple view and have established a central role of the lysosome in nutrient-dependent signal transduction. The degradative role of the lysosome and its newly discovered signaling functions are not in conflict but rather cooperate extensively to mediate fundamental cellular activities such as nutrient sensing, metabolic adaptation, and quality control of proteins and organelles. Moreover, lysosome-based signaling and degradation are subject to reciprocal regulation. Transcriptional programs of increasing complexity control the biogenesis, composition, and abundance of lysosomes and fine-tune their activity to match the evolving needs of the cell. Alterations in these essential activities are, not surprisingly, central to the pathophysiology of an ever-expanding spectrum of conditions, including storage disorders, neurodegenerative diseases, and cancer. Thus, unraveling the functions of this fascinating organelle will contribute to our understanding of the fundamental logic of metabolic organization and will point to novel therapeutic avenues in several human diseases.
Subject(s)
Lysosomes/metabolism , Animals , Disease , Exocytosis , Humans , Signal TransductionABSTRACT
As one of the two highly conserved cellular degradation systems, autophagy plays a critical role in regulation of protein, lipid, and organelle quality control and cellular homeostasis. This evolutionarily conserved pathway singles out intracellular substrates for elimination via encapsulation within a double-membrane vesicle and delivery to the lysosome for degradation. Multiple cancers disrupt normal regulation of autophagy and hijack its degradative ability to remodel their proteome, reprogram their metabolism, and adapt to environmental challenges, making the autophagy-lysosome system a prime target for anti-cancer interventions. Here, we discuss the roles of autophagy in tumor progression, including cancer-specific mechanisms of autophagy regulation and the contribution of tumor and host autophagy in metabolic regulation, immune evasion, and malignancy. We further discuss emerging proteomics-based approaches for systematic profiling of autophagosome-lysosome composition and contents. Together, these approaches are uncovering new features and functions of autophagy, leading to more effective strategies for targeting this pathway in cancer.
Subject(s)
Autophagosomes , Neoplasms , Autophagosomes/metabolism , Autophagy/physiology , Humans , Lysosomes/metabolism , Neoplasms/pathology , Quality ControlABSTRACT
Immune evasion is a major obstacle for cancer treatment. Common mechanisms of evasion include impaired antigen presentation caused by mutations or loss of heterozygosity of the major histocompatibility complex class I (MHC-I), which has been implicated in resistance to immune checkpoint blockade (ICB) therapy1-3. However, in pancreatic ductal adenocarcinoma (PDAC), which is resistant to most therapies including ICB4, mutations that cause loss of MHC-I are rarely found5 despite the frequent downregulation of MHC-I expression6-8. Here we show that, in PDAC, MHC-I molecules are selectively targeted for lysosomal degradation by an autophagy-dependent mechanism that involves the autophagy cargo receptor NBR1. PDAC cells display reduced expression of MHC-I at the cell surface and instead demonstrate predominant localization within autophagosomes and lysosomes. Notably, inhibition of autophagy restores surface levels of MHC-I and leads to improved antigen presentation, enhanced anti-tumour T cell responses and reduced tumour growth in syngeneic host mice. Accordingly, the anti-tumour effects of autophagy inhibition are reversed by depleting CD8+ T cells or reducing surface expression of MHC-I. Inhibition of autophagy, either genetically or pharmacologically with chloroquine, synergizes with dual ICB therapy (anti-PD1 and anti-CTLA4 antibodies), and leads to an enhanced anti-tumour immune response. Our findings demonstrate a role for enhanced autophagy or lysosome function in immune evasion by selective targeting of MHC-I molecules for degradation, and provide a rationale for the combination of autophagy inhibition and dual ICB therapy as a therapeutic strategy against PDAC.
Subject(s)
Adenocarcinoma/immunology , Autophagy/immunology , Carcinoma, Pancreatic Ductal/immunology , Histocompatibility Antigens Class I/immunology , Histocompatibility Antigens Class I/metabolism , Pancreatic Neoplasms/immunology , Tumor Escape/immunology , Adenocarcinoma/drug therapy , Adenocarcinoma/genetics , Adenocarcinoma/pathology , Animals , Antigen Presentation/drug effects , Antigen Presentation/immunology , Autophagy/drug effects , Autophagy/genetics , CD8-Positive T-Lymphocytes/drug effects , CD8-Positive T-Lymphocytes/immunology , Carcinoma, Pancreatic Ductal/drug therapy , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/pathology , Cell Cycle Checkpoints/drug effects , Cell Cycle Checkpoints/immunology , Cell Line, Tumor , Chloroquine/pharmacology , Female , Histocompatibility Antigens Class I/genetics , Humans , Intracellular Signaling Peptides and Proteins/metabolism , Lysosomes/drug effects , Lysosomes/metabolism , Male , Mice , Pancreatic Neoplasms/drug therapy , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Tumor Escape/drug effectsABSTRACT
Autophagy is a core molecular pathway for the preservation of cellular and organismal homeostasis. Pharmacological and genetic interventions impairing autophagy responses promote or aggravate disease in a plethora of experimental models. Consistently, mutations in autophagy-related processes cause severe human pathologies. Here, we review and discuss preclinical data linking autophagy dysfunction to the pathogenesis of major human disorders including cancer as well as cardiovascular, neurodegenerative, metabolic, pulmonary, renal, infectious, musculoskeletal, and ocular disorders.
Subject(s)
Autophagy , Disease Susceptibility , Animals , Autophagy/drug effects , Autophagy/genetics , Autophagy/immunology , Biomarkers , Gene Expression Regulation , Genetic Predisposition to Disease , Homeostasis , Host-Pathogen Interactions , Humans , Organ Specificity , Signal TransductionABSTRACT
The recent identification of several novel endocytic compartments has challenged our current understanding of the topological and functional organization of the endocytic pathway. Using quantitative single vesicle imaging and acute manipulation of phosphoinositides we show that APPL endosomes, which participate in growth factor receptor trafficking and signaling, represent an early endocytic intermediate common to a subset of clathrin derived endocytic vesicles and macropinosomes. Most APPL endosomes are precursors of classical PI3P positive endosomes, and PI3P plays a critical role in promoting this conversion. Depletion of PI3P causes a striking reversion of Rab5 positive endosomes to the APPL stage, and results in enhanced growth factor signaling. These findings reveal a surprising plasticity of the early endocytic pathway. Importantly, PI3P functions as a switch to dynamically regulate maturation and signaling of APPL endosomes.
Subject(s)
Endosomes/metabolism , Phosphatidylinositols/metabolism , Animals , COS Cells , Chlorocebus aethiops , Clathrin-Coated Vesicles/metabolism , Endocytosis , Signal TransductionABSTRACT
Activation of cellular stress response pathways to maintain metabolic homeostasis is emerging as a critical growth and survival mechanism in many cancers. The pathogenesis of pancreatic ductal adenocarcinoma (PDA) requires high levels of autophagy, a conserved self-degradative process. However, the regulatory circuits that activate autophagy and reprogram PDA cell metabolism are unknown. Here we show that autophagy induction in PDA occurs as part of a broader transcriptional program that coordinates activation of lysosome biogenesis and function, and nutrient scavenging, mediated by the MiT/TFE family of transcription factors. In human PDA cells, the MiT/TFE proteins--MITF, TFE3 and TFEB--are decoupled from regulatory mechanisms that control their cytoplasmic retention. Increased nuclear import in turn drives the expression of a coherent network of genes that induce high levels of lysosomal catabolic function essential for PDA growth. Unbiased global metabolite profiling reveals that MiT/TFE-dependent autophagy-lysosome activation is specifically required to maintain intracellular amino acid pools. These results identify the MiT/TFE proteins as master regulators of metabolic reprogramming in pancreatic cancer and demonstrate that transcriptional activation of clearance pathways converging on the lysosome is a novel hallmark of aggressive malignancy.
Subject(s)
Autophagy/genetics , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/metabolism , Gene Expression Regulation, Neoplastic , Lysosomes/metabolism , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/pathology , Transcription Factors/metabolism , Active Transport, Cell Nucleus , Amino Acids/metabolism , Animals , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Carcinoma, Pancreatic Ductal/pathology , Cell Line, Tumor , Energy Metabolism , Female , Heterografts , Homeostasis , Humans , Lysosomes/genetics , Mice , Microphthalmia-Associated Transcription Factor/metabolism , Neoplasm Transplantation , Pancreatic Neoplasms/genetics , Transcription, GeneticABSTRACT
Cancer cells have metabolic dependencies that distinguish them from their normal counterparts. Among these dependencies is an increased use of the amino acid glutamine to fuel anabolic processes. Indeed, the spectrum of glutamine-dependent tumours and the mechanisms whereby glutamine supports cancer metabolism remain areas of active investigation. Here we report the identification of a non-canonical pathway of glutamine use in human pancreatic ductal adenocarcinoma (PDAC) cells that is required for tumour growth. Whereas most cells use glutamate dehydrogenase (GLUD1) to convert glutamine-derived glutamate into α-ketoglutarate in the mitochondria to fuel the tricarboxylic acid cycle, PDAC relies on a distinct pathway in which glutamine-derived aspartate is transported into the cytoplasm where it can be converted into oxaloacetate by aspartate transaminase (GOT1). Subsequently, this oxaloacetate is converted into malate and then pyruvate, ostensibly increasing the NADPH/NADP(+) ratio which can potentially maintain the cellular redox state. Importantly, PDAC cells are strongly dependent on this series of reactions, as glutamine deprivation or genetic inhibition of any enzyme in this pathway leads to an increase in reactive oxygen species and a reduction in reduced glutathione. Moreover, knockdown of any component enzyme in this series of reactions also results in a pronounced suppression of PDAC growth in vitro and in vivo. Furthermore, we establish that the reprogramming of glutamine metabolism is mediated by oncogenic KRAS, the signature genetic alteration in PDAC, through the transcriptional upregulation and repression of key metabolic enzymes in this pathway. The essentiality of this pathway in PDAC and the fact that it is dispensable in normal cells may provide novel therapeutic approaches to treat these refractory tumours.
Subject(s)
Glutamine/metabolism , Metabolic Networks and Pathways , Oncogene Protein p21(ras)/metabolism , Pancreatic Neoplasms/metabolism , Pancreatic Neoplasms/pathology , Proto-Oncogene Proteins/metabolism , ras Proteins/metabolism , Adenocarcinoma/genetics , Adenocarcinoma/metabolism , Adenocarcinoma/pathology , Aspartate Aminotransferases/deficiency , Aspartate Aminotransferases/genetics , Aspartate Aminotransferases/metabolism , Cell Line, Tumor , Cell Proliferation , Citric Acid Cycle , Glutamate Dehydrogenase/metabolism , Homeostasis , Humans , Ketoglutaric Acids/metabolism , Oncogene Protein p21(ras)/genetics , Oncogenes/genetics , Oxidation-Reduction , Pancreatic Neoplasms/genetics , Proto-Oncogene Proteins/genetics , Proto-Oncogene Proteins p21(ras) , Reactive Oxygen Species/metabolism , ras Proteins/geneticsABSTRACT
Pancreatic ductal adenocarcinoma (PDA) is the most lethal of common human malignancies, with no truly effective therapies for advanced disease. Preclinical studies have suggested a therapeutic benefit of targeting the Hedgehog (Hh) signaling pathway, which is activated throughout the course of PDA progression by expression of Hh ligands in the neoplastic epithelium and paracrine response in the stromal fibroblasts. Clinical trials to test this possibility, however, have yielded disappointing results. To further investigate the role of Hh signaling in the formation of PDA and its precursor lesion, pancreatic intraepithelial neoplasia (PanIN), we examined the effects of genetic or pharmacologic inhibition of Hh pathway activity in three distinct genetically engineered mouse models and found that Hh pathway inhibition accelerates rather than delays progression of oncogenic Kras-driven disease. Notably, pharmacologic inhibition of Hh pathway activity affected the balance between epithelial and stromal elements, suppressing stromal desmoplasia but also causing accelerated growth of the PanIN epithelium. In striking contrast, pathway activation using a small molecule agonist caused stromal hyperplasia and reduced epithelial proliferation. These results indicate that stromal response to Hh signaling is protective against PDA and that pharmacologic activation of pathway response can slow tumorigenesis. Our results provide evidence for a restraining role of stroma in PDA progression, suggesting an explanation for the failure of Hh inhibitors in clinical trials and pointing to the possibility of a novel type of therapeutic intervention.
Subject(s)
Carcinoma, Pancreatic Ductal/metabolism , Hedgehog Proteins/metabolism , Pancreatic Neoplasms/metabolism , Proto-Oncogene Proteins p21(ras)/metabolism , Signal Transduction , Animals , Carcinoma, Pancreatic Ductal/drug therapy , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/pathology , Hedgehog Proteins/antagonists & inhibitors , Hedgehog Proteins/genetics , Humans , Mice , Mice, Knockout , Pancreatic Neoplasms/drug therapy , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Proto-Oncogene Proteins p21(ras)/geneticsABSTRACT
BACKGROUND: The best surgical approach for tumors of the proximal stomach remains controversial. For proximal gastrectomy (PG), the evidence regarding quality of life (QoL) and functional outcomes is controversial. Moreover, there are limited data from non-Asian settings. METHODS: All patients who underwent PG from September 2005 to July 2013 were identified from an institutional database. Demographic, perioperative and pathologic characteristics were retrieved. Symptom scores (0 = best/4 = worst) for reflux symptoms, dysphagia and validated QoL metrics (FACT scale, where a higher score is better) were assessed during early and late follow-up. Eligible patients for analysis were those with no evidence of recurrence. RESULTS: Of 465 upper gastrointestinal cancer resections, 50 were PG for adenocarcinoma (42; 84%), neuroendocrine carcinoma (5; 10%) or other pathologies (3; 6%). R0 resection was achieved in 44 (89.8%) of 49 patients with malignant tumors. Median lymph node collection was 32 (range 7-57). QoL scores did not differ from preoperative to early follow-up but increased compared to both at late follow-up [preoperative, 125 (interquartile range 105-140); early follow-up, 122.5 (97-142); late follow-up, 147 (132-159); p < 0.05]. At early and late follow-up, 9 (21.4%) of 42 and 10 (33.3%) of 30 patients reported reflux symptoms, but most were mild. Endoscopic signs of esophagitis were found in 7 (29%) of 24 patients, but only two of these reported reflux symptoms. Conversely only three of eight patients with reflux symptoms had esophagitis on endoscopy. CONCLUSIONS: Global QoL is not reduced early after PG, and increases compared to baseline at late follow-up. Although reflux symptoms are reported by a quarter of patients, most are mild, and there is little correlation with esophagitis. PG should remain a viable option in the management of proximal gastric tumors.
Subject(s)
Esophagogastric Junction/pathology , Esophagostomy , Gastrectomy , Gastrostomy , Postoperative Complications , Quality of Life , Stomach Neoplasms/pathology , Adenocarcinoma/pathology , Adenocarcinoma/surgery , Adult , Aged , Aged, 80 and over , Carcinoma, Neuroendocrine/pathology , Carcinoma, Neuroendocrine/surgery , Esophagogastric Junction/surgery , Female , Follow-Up Studies , Humans , Male , Middle Aged , Neoplasm Staging , Prognosis , Prospective Studies , Stomach Neoplasms/surgeryABSTRACT
The microphthalmia/transcription factor E (MiT/TFE) transcription factors (TFs; TFEB, TFE3, MITF, and TFEC) play a central role in cellular catabolism and quality control and are subject to extensive layers of regulation that influence their localization, stability, and activity. Recent studies have highlighted a broader role for these TFs in driving diverse stress-adaptation pathways, which manifest in a context- and tissue-dependent manner. Several human cancers upregulate the MiT/TFE factors to survive extreme fluctuations in nutrients, energy, and pharmacological challenges. Emerging data suggest that reduced activity of the MiT/TFE factors can also promote tumorigenesis. Here, we outline recent findings relating to novel mechanisms of regulation and activity of MiT/TFE proteins across some of the most aggressive human cancers.
Subject(s)
Microphthalmos , Neoplasms , Humans , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/genetics , Basic Helix-Loop-Helix Leucine Zipper Transcription Factors/metabolism , Microphthalmia-Associated Transcription Factor/genetics , Microphthalmia-Associated Transcription Factor/metabolism , Microphthalmos/metabolism , Lysosomes/metabolism , Neoplasms/genetics , Neoplasms/metabolismABSTRACT
Candidate cis-regulatory elements (cCREs) in microglia demonstrate the most substantial enrichment for Alzheimer's disease (AD) heritability compared to other brain cell types. However, whether and how these genome-wide association studies (GWAS) variants contribute to AD remain elusive. Here we prioritize 308 previously unreported AD risk variants at 181 cCREs by integrating genetic information with microglia-specific 3D epigenome annotation. We further establish the link between functional variants and target genes by single-cell CRISPRi screening in microglia. In addition, we show that AD variants exhibit allelic imbalance on target gene expression. In particular, rs7922621 is the effective variant in controlling TSPAN14 expression among other nominated variants in the same cCRE and exerts multiple physiological effects including reduced cell surface ADAM10 and altered soluble TREM2 (sTREM2) shedding. Our work represents a systematic approach to prioritize and characterize AD-associated variants and provides a roadmap for advancing genetic association to experimentally validated cell-type-specific phenotypes and mechanisms.
Subject(s)
Alzheimer Disease , Humans , Alzheimer Disease/genetics , Alzheimer Disease/metabolism , Microglia/metabolism , Genome-Wide Association Study , Cell Membrane/metabolism , PhenotypeSubject(s)
Cell Transformation, Neoplastic/metabolism , Cell Transformation, Neoplastic/pathology , NF-E2-Related Factor 2/metabolism , Oncogenes/genetics , Reactive Oxygen Species/metabolism , Adaptor Proteins, Signal Transducing/genetics , Adaptor Proteins, Signal Transducing/metabolism , Animals , Antioxidants/metabolism , Cell Transformation, Neoplastic/genetics , Cytoskeletal Proteins/genetics , Cytoskeletal Proteins/metabolism , Extracellular Signal-Regulated MAP Kinases/metabolism , Humans , Intracellular Signaling Peptides and Proteins/genetics , Intracellular Signaling Peptides and Proteins/metabolism , JNK Mitogen-Activated Protein Kinases/metabolism , Kelch-Like ECH-Associated Protein 1 , MAP Kinase Signaling System , Mice , Mitogen-Activated Protein Kinase Kinases/metabolism , NF-E2-Related Factor 2/genetics , Neoplasms/genetics , Neoplasms/metabolism , Neoplasms/pathologyABSTRACT
Lysosomes play major roles in growth regulation and catabolism and are recognized as critical mediators of cellular remodeling. An emerging theme is how the lysosome is itself subjected to extensive remodeling in order to perform specific tasks that meet the changing demands of the cell. Accordingly, lysosomes can sustain physical damage and undergo dramatic changes in composition following pathogen infection, accumulation of protein aggregates, or cellular transformation, necessitating dedicated pathways for their repair, remodeling, and restoration. In this review, we focus on emerging molecular mechanisms for piecemeal remodeling of lysosomal components and wholesale repair and discuss their implications in physiological and pathogenic challenges such as cancer, neurodegeneration, and pathogen infection.
Subject(s)
Lysosomes , Neoplasms , Humans , Lysosomes/metabolism , Neoplasms/pathologyABSTRACT
Lysosomes coordinate cellular metabolism and growth upon sensing of essential nutrients, including cholesterol. Through bioinformatic analysis of lysosomal proteomes, we identified lysosomal cholesterol signaling (LYCHOS, previously annotated as G protein-coupled receptor 155), a multidomain transmembrane protein that enables cholesterol-dependent activation of the master growth regulator, the protein kinase mechanistic target of rapamycin complex 1 (mTORC1). Cholesterol bound to the amino-terminal permease-like region of LYCHOS, and mutating this site impaired mTORC1 activation. At high cholesterol concentrations, LYCHOS bound to the GATOR1 complex, a guanosine triphosphatase (GTPase)-activating protein for the Rag GTPases, through a conserved cytoplasm-facing loop. By sequestering GATOR1, LYCHOS promotes cholesterol- and Rag-dependent recruitment of mTORC1 to lysosomes. Thus, LYCHOS functions in a lysosomal pathway for cholesterol sensing and couples cholesterol concentrations to mTORC1-dependent anabolic signaling.
Subject(s)
Cholesterol , Lysosomes , Mechanistic Target of Rapamycin Complex 1 , Receptors, G-Protein-Coupled , Cholesterol/metabolism , GTPase-Activating Proteins/metabolism , Humans , Lysosomes/metabolism , Mechanistic Target of Rapamycin Complex 1/metabolism , Monomeric GTP-Binding Proteins/metabolism , Proteome/metabolism , Receptors, G-Protein-Coupled/metabolismABSTRACT
The mechanisms underlying metabolic adaptation of pancreatic ductal adenocarcinoma (PDA) cells to pharmacologic inhibition of RAS-MAPK signaling are largely unknown. Using transcriptome and chromatin immunoprecipitation profiling of PDA cells treated with the MEK inhibitor (MEKi) trametinib, we identify transcriptional antagonism between c-MYC and the master transcription factors for lysosome gene expression, the MiT/TFE proteins. Under baseline conditions, c-MYC and MiT/TFE factors compete for binding to lysosome gene promoters to fine-tune gene expression. Treatment of PDA cells or patient organoids with MEKi leads to c-MYC downregulation and increased MiT/TFE-dependent lysosome biogenesis. Quantitative proteomics of immunopurified lysosomes uncovered reliance on ferritinophagy, the selective degradation of the iron storage complex ferritin, in MEKi-treated cells. Ferritinophagy promotes mitochondrial iron-sulfur cluster protein synthesis and enhanced mitochondrial respiration. Accordingly, suppressing iron utilization sensitizes PDA cells to MEKi, highlighting a critical and targetable reliance on lysosome-dependent iron supply during adaptation to KRAS-MAPK inhibition. SIGNIFICANCE: Reduced c-MYC levels following MAPK pathway suppression facilitate the upregulation of autophagy and lysosome biogenesis. Increased autophagy-lysosome activity is required for increased ferritinophagy-mediated iron supply, which supports mitochondrial respiration under therapy stress. Disruption of ferritinophagy synergizes with KRAS-MAPK inhibition and blocks PDA growth, thus highlighting a key targetable metabolic dependency. See related commentary by Jain and Amaravadi, p. 2023. See related article by Santana-Codina et al., p. 2180. This article is highlighted in the In This Issue feature, p. 2007.
Subject(s)
Carcinoma, Pancreatic Ductal , Iron-Sulfur Proteins , Pancreatic Neoplasms , Humans , Biological Availability , Carcinoma, Pancreatic Ductal/drug therapy , Carcinoma, Pancreatic Ductal/genetics , Carcinoma, Pancreatic Ductal/metabolism , Iron/metabolism , Iron/therapeutic use , Iron-Sulfur Proteins/metabolism , Iron-Sulfur Proteins/therapeutic use , Nuclear Receptor Coactivators/metabolism , Pancreatic Neoplasms/drug therapy , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/metabolism , Protein Kinase Inhibitors/pharmacology , Protein Kinase Inhibitors/therapeutic use , Proto-Oncogene Proteins p21(ras)/genetics , Proto-Oncogene Proteins p21(ras)/metabolism , Sulfur/metabolism , Sulfur/therapeutic use , Transcription Factors/metabolism , Pancreatic NeoplasmsABSTRACT
Today's cell biology could be considered a fusion of disciplines that blends advanced genetics, molecular biology, biochemistry, and engineering to answer fundamental as well as medically relevant scientific questions. Accordingly, our understanding of diseases is greatly aided by an existing vast knowledge base of fundamental cell biology. Gunter Blobel captured this concept when he said, "the tremendous acquisition of basic knowledge will allow a much more rational treatment of cancer, viral infection, degenerative disease and mental disease." In other words, without cell biology can we truly understand, prevent, or effectively treat a disease?
Subject(s)
Cell Biology , Lysosomes/pathology , Neoplasms/pathology , Animals , Autophagy/genetics , Humans , Mice , Neoplasms/drug therapy , Neoplasms, Experimental/genetics , Neoplasms, Experimental/pathologyABSTRACT
Oncogenes can alter metabolism by changing the balance between anabolic and catabolic processes. However, how oncogenes regulate tumor cell biomass remains poorly understood. Using isogenic MCF10A cells transformed with nine different oncogenes, we show that specific oncogenes reduce the biomass of cancer cells by promoting extracellular vesicle (EV) release. While MYC and AURKB elicited the highest number of EVs, each oncogene selectively altered the protein composition of released EVs. Likewise, oncogenes alter secreted miRNAs. MYC-overexpressing cells require ceramide, whereas AURKB requires ESCRT to release high levels of EVs. We identify an inverse relationship between MYC upregulation and activation of the RAS/MEK/ERK signaling pathway for regulating EV release in some tumor cells. Finally, lysosome genes and activity are downregulated in the context of MYC and AURKB, suggesting that cellular contents, instead of being degraded, were released via EVs. Thus, oncogene-mediated biomass regulation via differential EV release is a new metabolic phenotype.
Subject(s)
Aurora Kinase B/genetics , Extracellular Vesicles/metabolism , Oncogenes/genetics , Proto-Oncogene Proteins c-myc/genetics , Energy Metabolism/genetics , Extracellular Vesicles/genetics , Gene Expression Regulation, Neoplastic , Genes, ras/genetics , Humans , Lysosomes/genetics , MAP Kinase Kinase Kinases/genetics , MAP Kinase Signaling System/genetics , Metabolism/genetics , Signal Transduction/geneticsABSTRACT
Lysosomes must maintain the integrity of their limiting membrane to ensure efficient fusion with incoming organelles and degradation of substrates within their lumen. Pancreatic cancer cells upregulate lysosomal biogenesis to enhance nutrient recycling and stress resistance, but it is unknown whether dedicated programmes for maintaining the integrity of the lysosome membrane facilitate pancreatic cancer growth. Using proteomic-based organelle profiling, we identify the Ferlin family plasma membrane repair factor Myoferlin as selectively and highly enriched on the membrane of pancreatic cancer lysosomes. Mechanistically, lysosomal localization of Myoferlin is necessary and sufficient for the maintenance of lysosome health and provides an early acting protective system against membrane damage that is independent of the endosomal sorting complex required for transport (ESCRT)-mediated repair network. Myoferlin is upregulated in human pancreatic cancer, predicts poor survival and its ablation severely impairs lysosome function and tumour growth in vivo. Thus, retargeting of plasma membrane repair factors enhances the pro-oncogenic activities of the lysosome.
Subject(s)
Biomarkers, Tumor/metabolism , Calcium-Binding Proteins/metabolism , Cell Proliferation , Intracellular Membranes/metabolism , Lysosomes/metabolism , Membrane Proteins/metabolism , Muscle Proteins/metabolism , Pancreatic Neoplasms/metabolism , Animals , Biomarkers, Tumor/genetics , Calcium-Binding Proteins/genetics , Cell Line, Tumor , Gene Expression Regulation, Neoplastic , Humans , Intracellular Membranes/pathology , Lysosomes/genetics , Lysosomes/pathology , Membrane Proteins/genetics , Mice, Inbred C57BL , Mice, Transgenic , Muscle Proteins/genetics , Pancreatic Neoplasms/genetics , Pancreatic Neoplasms/pathology , Prognosis , Signal Transduction , Tumor BurdenABSTRACT
Lysosomes promote cellular homeostasis through macromolecular hydrolysis within their lumen and metabolic signaling by the mTORC1 kinase on their limiting membranes. Both hydrolytic and signaling functions require precise regulation of lysosomal cholesterol content. In Niemann-Pick type C (NPC), loss of the cholesterol exporter, NPC1, causes cholesterol accumulation within lysosomes, leading to mTORC1 hyperactivation, disrupted mitochondrial function, and neurodegeneration. The compositional and functional alterations in NPC lysosomes and nature of aberrant cholesterol-mTORC1 signaling contribution to organelle pathogenesis are not understood. Through proteomic profiling of NPC lysosomes, we find pronounced proteolytic impairment compounded with hydrolase depletion, enhanced membrane damage, and defective mitophagy. Genetic and pharmacologic mTORC1 inhibition restores lysosomal proteolysis without correcting cholesterol storage, implicating aberrant mTORC1 as a pathogenic driver downstream of cholesterol accumulation. Consistently, mTORC1 inhibition ameliorates mitochondrial dysfunction in a neuronal model of NPC. Thus, cholesterol-mTORC1 signaling controls organelle homeostasis and is a targetable pathway in NPC.