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Defining the Function of RME-8 in Endosomal Regulation During Health and Disease

定义 RME-8 在健康和疾病期间内体调节中的功能

基本信息

批准号：
10334441
负责人：
Qian Cai
金额：
$ 33.79万
依托单位：
RUTGERS, THE STATE UNIV OF N.J.
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-02-01 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10334441
关键词：
Acute Affect Alleles Animals Architecture Area Auxins Axon Axonal Transport Binding Biological Assay Biological Process C-terminal Caenorhabditis elegans Caliber Cell physiology Cells Clathrin Clathrin Heavy Chains Complex DNA Data Disease Dynein ATPase Endosomes Equilibrium Essential Tremor Fluorescence Functional disorder Genetic Genetic Techniques Gilles de la Tourette syndrome Goals Golgi Apparatus Health Human Impairment In Vitro Individual Life Link Lysosomes Maintenance Mammals Membrane Membrane Microdomains Methods Modeling Molecular Molecular Chaperones Molecular Genetics Motor Movement Mus Nematoda Neurodegenerative Disorders Neurons Organelles Organism Parkinson Disease Pathway interactions Patients Peripheral Point Mutation Process Proteins Recycling Regulation Repression Research Resolution Role Sorting - Cell Movement System Techniques Tertiary Protein Structure Testing Time Transmembrane Transport Variant Visible Radiation Visualization Work cell motility combat dynactin experimental study in vivo in vivo Model insight light microscopy mutant nervous system disorder neuronal cell body novel prevent retrograde transport scaffold therapeutic target virtual

项目摘要

Our long-term goal in this proposal is to understand how endosomes differentially balance opposing activities (recycling versus degradation) within a single endosome, especially with respect to regulation by the only known endosomal DNA-J domain protein RME-8. We also seek to understand how RME-8 contributes to neurodegenerative disease. To gain new insights into the mechanisms that balance opposing endosomal functions, we have pioneered the use of the coelomocyte cells of the nematode C. elegans for the direct visualization of endosomal microdomains, and then applied powerful molecular genetic techniques to identify and decipher metazoan- specific mechanisms that control endosome function and functional microdomain separation. We then extend this work into mammalian systems. In previous studies, we identified the J-domain protein RME-8 as a metazoan specific endosome regulator. We further showed that RME-8 functions with Retromer component SNX-1/Snx1, and the chaperone Hsc70, to promote the recycling of retromer cargo in C. elegans and mammals. Most recently, we showed that RME-8 allows Retromer to negatively regulate ESCRT assembly on endosomes, a process required to prevent mixing of recycling and degradative microdomains. Here we propose to define how SNX-1 regulates RME-8, testing a de-repression model for RME-8 activation. We then focus on mechanistic questions of how RME-8 inhibits ESCRT microdomain expansion through the endosomal flat clathrin lattice and newly identified RME-8 interacting proteins. Finally, we use primary mouse neurons and in vivo C. elegans analysis to decipher neuronal functions of RME-8 as revealed by analysis of a familial Parkinson’s associated allele of RME-8. We propose to define how the disease allele affects RME-8 function in endosomal recycling and microdomain maintenance, and determine how RME-8 regulates long-distance axonal transport of endosomes, an additional transport step in neurons required for endocytic cargo sorting. A better mechanistic understanding of these regulatory mechanisms of endosomes will be profoundly important in identifying therapeutic targets to combat diseases associated with endolysosomal dysfunction.

我们在这项提议中的长期目标是了解内体是如何差异化地平衡单个内体内的相反活性（再循环与降解），特别是关于唯一已知的内体DNA-J结构域的调节蛋白质RME-8。我们还试图了解RME-8如何有助于神经退行性疾病为了获得新的见解，相反的内体功能，我们已经率先使用体腔细胞的细胞，线虫C. elegans用于直接可视化内体微结构域，和然后应用强大的分子遗传技术来识别和破译后生动物- 控制内体功能和功能微区的特定机制分居然后，我们将这项工作扩展到哺乳动物系统。在以前的研究中，我们鉴定了J结构域蛋白RME-8作为后生动物特异性内体调节剂。我们进一步表明RME-8与Retromer组分SNX-1/Snx 1一起起作用，伴侣蛋白Hsc 70，以促进C.线虫和哺乳动物最近，我们发现RME-8允许Retromer负调节 ESCRT在内体上组装，这是防止再循环和再利用的混合所需的过程。降解微区。在这里，我们建议定义SNX-1如何调节RME-8，测试RME-8激活的去阻遏模型。然后我们专注于机械关于RME-8如何抑制ESCRT微区扩展的问题，内体扁平网格蛋白晶格和新鉴定的RME-8相互作用蛋白。最后，我们使用原代小鼠神经元和体内C.用elegans分析来解读神经元 RME-8的功能，如通过分析家族性帕金森病相关等位基因所揭示的， RME-8。我们建议确定疾病等位基因如何影响RME-8功能，内体再循环和微区维护，并确定如何RME-8 调节核内体的长距离轴突运输，这是核内体中的一个额外的运输步骤。内吞货物分选所需的神经元。更好的机械理解内体的这些调节机制在识别治疗靶点以对抗与内溶酶体功能障碍相关的疾病。