Dissecting new mechanisms of lysosome quality control in health and disease

剖析健康和疾病中溶酶体质量控制的新机制

• 批准号: 10370440
• 负责人: Rushika Miriam Perera
• 金额: $ 36.2万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2026-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10370440
• 关键词: Acute; Affinity; Aging; Architecture; Autophagocytosis; Binding; Biochemical; Biochemistry; Biogenesis; Biological Assay; C-terminal; C2 Domain; Calcium; Catabolism; Cell Proliferation; Cell membrane; Cell physiology; Cells; Characteristics; Chemicals; Coupled; Cytoplasm; DYSF gene; Data; Defect; Degenerative Disorder; Dependence; Digestion; Disease; Disease Progression; Dissection; Ensure; Family; Family member; Fluorescent Probes; Goals; Golgi Apparatus; Growth; Health; Homeostasis; Impairment; In Vitro; Light; Lipid Bilayers; Lysosomes; Maintenance; Malignant Neoplasms; Malignant neoplasm of pancreas; Mass Spectrum Analysis; Measurement; Measures; Mechanical Stress; Mediating; Membrane; Membrane Proteins; Metabolic; Metabolism; Mitochondria; Morphology; Muscular Dystrophies; Mutation; Myoblasts; Normal Cell; Nutrient; Organelles; Pancreatic Ductal Adenocarcinoma; Pathogenesis; Pathway interactions; Phospholipids; Phosphotransferases; Process; Property; Protein Family; Proteins; Proteomics; Quality Control; Reaction; Recycling; Resistance; Role; Route; Signal Transduction; Skeletal Muscle; Stress; Testing; Transmembrane Domain; base; cell type; comparative; detection of nutrient; in vivo; insight; lysosome membrane; member; metabolomics; mutant; novel; pancreatic ductal adenocarcinoma cell; preservation; programs; repaired; resilience; response; trafficking; transcription factor; tumor growth

PROJECT SUMMARY Lysosomes function as critical nodes for macromolecular recycling, metabolic rewiring, and pro-growth signaling in cells. Accordingly, defects in lysosome function underlie degenerative diseases and aging while hyperactivation of lysosomes are associated with cancer. Prior studies have shown that highly aggressive Pancreatic ductal adenocarcinoma (PDA) cells upregulate lysosome biogenesis and activity to facilitate degradation, clearance and recycling of incoming cargo material delivered by increased rates of autophagy and macropinocytosis. Whether qualitative differences endow PDA lysosomes with unique structural and functional properties to cope with a higher demand for substrate clearance remains unknown. To answer this question, we have conducted the first comparative proteomics analysis of lysosomes isolated from PDA versus normal cells and have identified members of the Ferlin family of membrane repair factors, Myoferlin and Dysferlin, as selectively enriched on the membrane of PDA lysosomes. We propose that Ferlin proteins confer increased protection against lysosomal membrane stress in PDA cells. Ferlin proteins are normally localized on the plasma membrane of cell types subjected to heightened mechanical stress, such as skeletal muscle, where they facilitate repair of the lipid bilayer. Accordingly, mutations in DYSF are associated with two forms of muscular dystrophy whereby impaired membrane resealing compromises myoblast maturation, fusion and plasma membrane repair. Therefore, we hypothesize that PDA cells hijack and repurpose Ferlin proteins at the lysosome membrane to protect the integrity of this organelle. In support of this hypothesis, our preliminary findings show that PDA lysosomes are more resistant to acute chemically induced membrane permeabilization relative to normal cells. Mechanistically, lysosome localization of MYOF is necessary and sufficient for maintenance of lysosome quality control and its suppression leads to profound defects in lysosome morphology, PDA cell proliferation and in vivo tumor growth. The goal of this study is to investigate how MYOF functions to protect the lysosome membrane in mechanistic detail and to determine the impact of blocking lysosome quality control on cellular metabolism, pro-growth signaling and disease progression. In summary, our discovery and proposed studies will be the first to determine a novel function for Ferlin proteins at the lysosome membrane and provide insight into how enhanced lysosome quality control regulates cellular homeostasis and disease pathogenesis.

项目总结 溶酶体是大分子循环、代谢重联和促进生长的关键节点。 细胞中的信号。因此，溶酶体功能缺陷是退行性疾病和衰老的基础 溶酶体的过度激活与癌症有关。先前的研究表明，高度攻击性的 胰腺导管腺癌(PDA)细胞上调溶酶体的生物发生和活性促进 通过提高自噬和自噬速度运送的入境货物材料的降解、清关和回收 巨噬细胞增多症。质的差异是否赋予PDA溶酶体独特的结构和功能 对于衬底清晰度的更高要求，其性能尚不清楚。为了回答这个问题，我们 首次对从pda和正常细胞分离的溶酶体进行了比较蛋白质组学分析。 并已确定膜修复因子Ferlin家族的成员Myoferlin和deferlin为 选择性地富集在PDA溶酶体膜上。我们认为费林蛋白赋予 增强PDA细胞对溶酶体膜应激的保护作用。 Ferlin蛋白通常定位于细胞类型的质膜上 机械应力，如骨骼肌，在那里它们有助于脂质双层的修复。相应地，突变 在DYSF中，DYSF与两种形式的肌营养不良有关，膜重新封闭受损 影响成肌细胞成熟、融合和质膜修复。因此，我们假设掌上电脑 细胞劫持并重新利用溶酶体膜上的费林蛋白，以保护这个细胞器的完整性。在……里面 支持这一假设，我们的初步发现表明，PDA溶酶体对急性 化学诱导的相对正常细胞的膜通透性。从机制上讲，溶酶体定位 MYOF是维持溶酶体质量控制的必要条件和充分条件，其抑制导致 在溶酶体形态、PDA细胞增殖和体内肿瘤生长方面存在严重缺陷。这样做的目的是 研究的目的是研究MYOF如何从机械细节上保护溶酶体膜，并 确定阻断溶酶体质量控制对细胞代谢、促生长信号和 疾病的发展。总而言之，我们的发现和拟议的研究将是第一次确定一部小说 FERLIN蛋白在溶酶体膜上的功能，并提供了对如何提高溶酶体质量的洞察 CONTROL调节细胞内稳态和疾病的发病机制。