Autophagy and LC3-associated phagocytosis in intestinal epithelial cells

肠上皮细胞中的自噬和 LC3 相关的吞噬作用

基本信息

批准号：
10538801
负责人：
Jean M Wilson
金额：
$ 45.09万
依托单位：
UNIVERSITY OF ARIZONA
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-08-01 至 2027-05-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10538801
关键词：
Autophagocytosis Autophagosome Bacteria Biogenesis Cell physiology Cells Crohn&apos s disease Data Development Endosomes Enterocytes Epithelial Epithelial Cells Functional disorder Genes Genetic Transcription Homeostasis Impairment Infection Inflammatory Bowel Diseases Innate Immune Response Integral Membrane Protein Intestinal Diseases Intestines Invaded Knock-out Knowledge Link Lysosomes Maintenance Mediating Mediator of activation protein Membrane Messenger RNA Metabolism Mitochondria Molecular Monomeric GTP-Binding Proteins Morphology Mucosal Immunity Mucous Membrane Names Natural Immunity Organoids Pathway interactions Patients Persons Phagocytes Phagocytosis Phagosomes Predisposition Process Proteins Regulation Risk Role Site Structure Testing Tight Junctions Tubular formation Variant Vesicle Virus Work cell type endotubin enteric pathogen experimental study insight intestinal epithelium intestinal homeostasis knock-down mutant pathogen prevent recruit risk variant trafficking transcriptome sequencing uptake

项目摘要

Abstract The intestinal epithelium is vital to maintain the barrier between the body and the outside world, and membrane trafficking is essential for both the development and maintenance of this barrier. Abnormal membrane trafficking can result compromised barrier leading to intestinal disease, including inflammatory bowel disease. Autophagy is a specialized membrane trafficking process that allows cells to respond to changes in metabolism. In addition, autophagy is important for maintenance of the epithelial barrier and the innate immune response, mediating the isolation and degradation of intracellular pathogens. Furthermore, LC3-associated phagocytosis (LAP) is important for uptake and degradation of pathogens, and dysfunction of this pathway is associated with hyperinflammation. Importantly, variants of proteins in the autophagic and LAP pathways have been linked to increased susceptibility to Inflammatory Bowel Disease (IBD), and particularly Crohn’s disease, although the molecular mechanisms that underlie this connection remain incompletely understood. MAMDC4 is an integral membrane protein that localizes to endosomes of the intestinal epithelium. Deletion of MAMDC4 compromises intestinal enterocyte morphology, and RNAseq studies have indicated that MAMDC4 is down- regulated in IBD. In previous work, we found that MAMDC4 interacts with the small GTPase Rab14. In professional phagocytes, Rab14 is recruited to phagosomes and prevents phagosome-lysosome fusion but its role in the intestinal epithelium is unknown. In our preliminary data, we show that Rab 14 is present on autophagosomes and both Rab14 and MAMDC4 are present on membranes containing invading bacteria. Furthermore, deletion of MAMDC4 results in accumulation of autophagy and lysosomal markers on tubular membranes. These results suggest that MAMDC4 and Rab14 act in a molecular network to maintain mucosal immunity through control of autophagy or LAP. In this proposal we will use intestinal epithelial cells in culture, organoid culture, and patient-derived organoids to define the role of autophagy and/or LAP in intestinal homeostasis.

抽象的肠上皮对于维持身体与外界之间的屏障至关重要，肠膜运输对于这种异常膜的形成和维持至关重要。贩运可能导致屏障受损，导致肠道疾病，包括炎症性肠病。自噬是一种特殊的膜运输过程，使细胞能够对细胞膜的变化做出反应此外，自噬对于维持上皮屏障和先天免疫也很重要。反应，介导细胞内病原体的分离和降解此外，LC3 相关。吞噬作用（LAP）对于病原体的摄取和降解很重要，该途径的功能障碍重要的是，自噬和 LAP 途径中的蛋白质变异与过度炎症有关。与炎症性肠病（IBD），特别是克罗恩病的易感性增加有关，尽管 MAMDC4 的分子机制尚不完全清楚。一种定位于肠上皮内体的整合膜蛋白 MAMDC4 缺失。损害肠道肠细胞形态，RNAseq 研究表明 MAMDC4 下调在之前的工作中，我们发现 MAMDC4 与小 GTPase Rab14 相互作用。作为专业吞噬细胞，Rab14 被招募到吞噬体并阻止吞噬体-溶酶体融合，但其在我们的初步数据中，我们表明 Rab 14 存在于肠上皮细胞中。自噬体以及 Rab14 和 MAMDC4 都存在于含有入侵细菌的膜上。此外，MAMDC4 的缺失会导致肾小管上自噬和溶酶体标记物的积累。这些结果表明 MAMDC4 和 Rab14 在分子网络中发挥作用以维持粘膜。通过控制自噬或 LAP 来实现免疫在本提案中，我们将在培养物中使用肠上皮细胞，类器官培养和患者来源的类器官，以确定自噬和/或 LAP 在肠道中的作用体内平衡。