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

基本信息

批准号：
10594038
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
项目类别：
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/10594038
关键词：
Acceleration Acute Affinity Aging Architecture Autophagocytosis Binding Biochemical Biochemistry Biogenesis Biological Assay C-terminal C2 Domain Calcium Catabolism Cell Membrane Permeability Cell Proliferation Cell membrane Cell physiology Cells Characteristics Chemicals Coupled Cytoplasm DYSF gene Data Dedications Defect Degenerative Disorder Dependence Digestion Disease Disease Progression Dissection Endowment Ensure Family Family member Fluorescent Probes Goals Golgi Apparatus Growth Health Homeostasis Impairment In Vitro 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 cell type comparative coping detection of nutrient in vivo insight lysosome membrane member metabolomics mutant novel pancreatic ductal adenocarcinoma cell preservation programs repaired resilience response seal trafficking transcription factor tumor growth

项目摘要

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.

项目概要溶酶体作为大分子回收、代谢重连和促生长的关键节点发挥作用细胞内的信号传导。因此，溶酶体功能缺陷是退行性疾病和衰老的基础，而溶酶体的过度激活与癌症有关。先前的研究表明，具有高度攻击性的胰腺导管腺癌 (PDA) 细胞上调溶酶体生物合成和活性，以促进通过提高自噬率来降解、清除和回收传入的货物材料巨胞饮作用。质的差异是否赋予PDA溶酶体独特的结构和功能满足更高的基板间隙要求的特性仍然未知。为了回答这个问题，我们对从 PDA 与正常细胞中分离的溶酶体进行了首次比较蛋白质组学分析并确定了膜修复因子 Ferlin 家族的成员 Myoferlin 和 Dysferlin，如选择性富集于PDA溶酶体膜上。我们建议 Ferlin 蛋白赋予增强对 PDA 细胞溶酶体膜应激的保护。 Ferlin 蛋白通常定位于受到高度刺激的细胞类型的质膜上。机械应力，例如骨骼肌，它们促进脂质双层的修复。因此，突变 DYSF 中的 DYSF 与两种形式的肌营养不良症相关，其中膜重新密封受损损害成肌细胞成熟、融合和质膜修复。因此，我们假设 PDA 细胞在溶酶体膜上劫持并重新利用 Ferlin 蛋白，以保护该细胞器的完整性。在为了支持这一假设，我们的初步研究结果表明 PDA 溶酶体对急性相对于正常细胞，化学诱导的膜透化。从机制上讲，溶酶体定位 MYOF 的存在对于维持溶酶体质量控制是必要且充分的，其抑制会导致溶酶体形态、PDA 细胞增殖和体内肿瘤生长的严重缺陷。此举的目标研究目的是研究 MYOF 如何发挥保护溶酶体膜的机制细节并确定阻断溶酶体质量控制对细胞代谢、促生长信号传导和疾病进展。总之，我们的发现和提出的研究将首先确定一种新颖的 Ferlin 蛋白在溶酶体膜上的功能，并提供如何增强溶酶体质量的见解控制调节细胞稳态和疾病发病机制。