DISSECTION OF SARM1-INDUCED AXON DEGENERATION AND CELL DEATH

SARM1 诱导的轴突变性和细胞死亡的剖析

基本信息

批准号：
9058619
负责人：
Aaron Diantonio
金额：
$ 36.46万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-01 至 2019-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9058619
关键词：
26S proteasome Adaptor Signaling Protein Adenosine Diphosphate Ribose Alzheimer&apos s Disease Apoptosis Automobile Driving Axon Biochemical Biological Assay Brain Cell Death Cells Cessation of life Consumption Data Dimerization Disease Dissection Drosophila genus Enzymes Event Genetic Goals Health High Pressure Liquid Chromatography Injury Knowledge Label Lead Link Massive Parallel Sequencing Mediating Metabolic Metabolic stress Methods Modeling Molecular Molecular Analysis Mus Mutate Neurodegenerative Disorders Neuronal Injury Neurons Parkinson Disease Pathway interactions Peptides Pharmaceutical Preparations Play Process Proteins Reagent Research Role SAM Domain Sequence Analysis Series Signal Pathway Signal Transduction Sirtuins Site Site-Directed Mutagenesis Specificity Stroke Structural Models Surface System Tankyrase Tertiary Protein Structure Testing Therapeutic Agents Toll-like receptors Traumatic Brain Injury Virus Diseases Western Blotting axonal degeneration candidate validation cell injury design injured knock-down loss of function loss of function mutation mitochondrial dysfunction multicatalytic endopeptidase complex mutant nervous system disorder neuron loss novel novel therapeutics overexpression prevent programs protein function receptor research study small hairpin RNA therapeutic target tool treatment strategy

项目摘要

DESCRIPTION (provided by applicant): Neurodegenerative disorders as well as traumatic and ischemic injuries to the brain are characterized by neuronal cell death and axonal degeneration. Programmed cell death and axonal degenerative are distinct self-destructive programs that are invoked to eliminate damaged cells and/or axons; however, they are activated inappropriately in many neurological disorders. While the components of pathways leading to cell death are relatively well characterized, the mechanisms involved in axonal degeneration are largely unknown. Studies of the wlds mouse revealed that overexpression of Nmnat enzymes, which synthesize NAD, can prevent axonal degeneration and, more recently, a toll-like receptor adaptor protein called Sarm1 was discovered to be an important component of the intrinsic axon degeneration program. Additional studies, including from our own labs, show that Sarm1 can also promote cell death in neurons and other cells. Indeed, loss of Sarm1 protects neurons from metabolic stress and mitochondrial dysfunction. Together, these breakthroughs show that Sarm1 drives a general cell destruction program that we term sarmoptosis. Our molecular analysis of Sarm1 shows that the SAM domains are necessary for its multimerization, whereas the TIR domain is required for its ability to activate cell death and axonal degeneration. To study molecular aspects of sarmoptosis, we have developed a variety of tools including a regulable Sarm1 TIR domain dimerization system that allows us to trigger cell death or, using compartmentalized chambers, axonal degeneration in a controlled fashion. It is our goal to understand sarmoptosis so that therapeutic agents can be devised to block this process, as this could be a useful method for treating many neurological disorders. In this proposal, we outline experiments that utilize these reagents to define the molecular pathways engaged by Sarm1 to promote cell destruction. First, we will identify structural motifs in the Sarm1 TIR domain that activate cell destruction. We will identify these key functional residues by analyzing a series of site-directed TIR domain mutants. Second, we will identify the enzyme(s) involved in mediating the NAD depletion that occurs after Sarm1 activation. Third, we will identify proteins that function downstream in this destructive pathway using a suppressor screen. Mechanistic studies of identified suppressors will be performed to characterize their role in sarmoptosis.

描述（由申请人提供）：神经退行性疾病以及大脑的创伤性和缺血性损伤的特征是神经元细胞死亡和轴突变性。程序性细胞死亡和轴突退化是不同的自我毁灭性程序，可以消除受损的细胞和/或轴突；但是，它们在许多神经系统疾病中被不适当地激活。虽然导致细胞死亡的途径的成分相对很好地表征，但与轴突变性有关的机制在很大程度上尚不清楚。对WLDS小鼠的研究表明，合成NAD的NMNAT酶的过表达可以防止轴突变性，并且最近发现了一种称为SARM1的Toll-like受体适配器蛋白是固有轴突退化程序的重要组成部分。包括我们自己的实验室在内的其他研究表明，SARM1还可以促进神经元和其他细胞中的细胞死亡。实际上，SARM1的丧失可以保护神经元免受代谢胁迫和线粒体功能障碍的侵害。这些突破共同表明，SARM1驱动了我们称为Sarmoptosis的一般细胞破坏程序。我们对SARM1的分子分析表明，SAM域对于其多聚化是必需的，而TIR结构域激活细胞死亡和轴突变性的能力是必需的。为了研究Sarmoptosis的分子方面，我们开发了各种工具，包括可调节的SARM1 TIR域二聚化系统，该系统使我们能够触发细胞死亡或使用分室化腔室，以受控的方式轴突变性。我们的目标是理解肌凋亡，以便可以设计治疗剂来阻止这一过程，因为这可能是治疗许多神经系统疾病的有用方法。在此提案中，我们概述了利用这些试剂来定义SARM1参与的分子途径以促进细胞破坏的实验。首先，我们将确定激活细胞破坏的SARM1 TIR结构域中的结构基序。我们将通过分析一系列位置定向的TIR结构域突变体来识别这些关键功能残基。其次，我们将确定介导SARM1激活后发生的NAD耗竭的酶。第三，我们将使用抑制器屏幕识别在这种破坏性途径中下游功能的蛋白质。将对已识别抑制剂进行机械研究，以表征其在刺凋亡中的作用。