Global Identification of Mammalian Synaptic Caspase Targets

哺乳动物突触 Caspase 靶标的全局识别

• 批准号: 8302120
• 负责人: James William Mandell
• 金额: $ 23.1万
• 依托单位: UNIVERSITY OF VIRGINIA
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-04-01 至 2014-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8302120
• 关键词: Affinity; Alzheimer' s Disease; Amyloid beta-Protein Precursor; Apoptosis; Apoptotic; Aspartate; Avidin; Biochemical; Biological; Biological Markers; Biotinylation; Brain; Brain region; Caspase; Cataloging; Catalogs; Cells; Cerebrospinal Fluid; Cessation of life; Cleaved cell; Complex; Computer Simulation; Cysteine; Databases; Development; Diagnosis; Disease; Enzymes; Ethanol toxicity; Event; Excision; Family; Functional disorder; Future; Homeostasis; Human; Huntington Disease; Knowledge; Label; Lead; Mass Spectrum Analysis; Mediating; Methods; Modeling; Molecular; Mus; N-terminal; Neurodegenerative Disorders; Neurons; Organism; Parkinson Disease; Patient Monitoring; Peptide Hydrolases; Peptides; Phagocytes; Population; Preparation; Process; Proteins; Proteome; Proteomics; Publishing; Regulation; Reporting; Research; Role; Sampling; Serum; Site; Stroke; Synapses; Synaptic Transmission; Synaptic plasticity; Synaptosomes; Trauma; Validation; Western Blotting; Work; brain tissue; human Huntingtin protein; in vivo Model; insight; mouse model; nervous system disorder; neurofibrillary tangle formation; neuron apoptosis; novel; protein complex; subtiligase; tau Proteins

DESCRIPTION (provided by applicant): Apoptosis, or programmed cell death, is a feature of all multicellular organisms and has critical roles in development, homeostasis, and disease. This intensely studied process is characterized by and requires the activation of a family of proteolytic enzymes known as caspases (cysteine-dependent aspartate-directed proteases). Caspases cleave a large number of specific target proteins, orchestrating the orderly transformation of intact cells into fragments that may be removed from the body by phagocytic cells. Recent evidence points to non-apoptotic roles for caspases in the brain, including the regulation of synaptic plasticity. Caspase activation was recently shown to precede tangle formation in a mouse model of Alzheimer's disease. However, surprisingly little is known about the global scope of neuronal synaptic proteins targeted by caspases, the specific sites of cleavage, and mechanisms by which caspase- mediated cleavages alter neuronal function. This proposal will use an N-terminus labeling approach to positively select cleaved proteins from the complex protein content of synaptosomes or brain lysates, allowing systematic identification caspase targets in synaptosomal preparations as well as in discrete brain regions and models of neuronal apoptotic induction. The analysis will not only identify the protein substrates of caspases in synapses, but will also reveal the precise site of proteolytic cleavage, providing immediate molecular insight into the function of the cleavage event. The information derived from this project will not only stimulate further research on the functional roles of caspase cleavage of synaptic targets, but will also provide immediate candidate biomarkers for human neurological disorders, including trauma, stroke, and neurodegenerative diseases. PUBLIC HEALTH RELEVANCE: Apoptosis, or programmed cell death, as well as the elimination of excess neuronal connections (synapses) is essential for normal brain development. However, the same process may be abnormally re-activated in neurodegenerative disorders such as Alzheimer's Disease. This proposal will use state-of-the-art proteomics methods to globally identify brain synaptic proteins that are cleaved by apoptotic enzymes called caspases. The information derived should provide immediate candidate biomarkers for human neurodegenerative diseases that could help in the diagnosis and monitoring of patients.

描述（由申请人提供）：细胞凋亡或程序性细胞死亡是所有多细胞生物的一个特征，在发育、体内平衡和疾病中起着关键作用。这一被深入研究的过程的特点是需要被称为半胱氨酸依赖的天冬氨酸导向蛋白酶（半胱氨酸依赖的天冬氨酸导向蛋白酶）的蛋白水解酶家族的激活。半胱天冬酶切割大量特定的靶蛋白，将完整细胞有序地转化为可被吞噬细胞从体内清除的片段。最近的证据指出半胱天冬酶在大脑中的非凋亡作用，包括调节突触可塑性。在阿尔茨海默病的小鼠模型中，最近发现Caspase的激活先于缠结的形成。然而，令人惊讶的是，关于半胱天冬酶靶向的神经元突触蛋白的整体范围，切割的特定位点以及半胱天冬酶介导的切割改变神经元功能的机制知之甚少。该提案将使用n端标记方法从突触体或脑裂解物的复杂蛋白含量中积极选择裂解蛋白，从而允许系统地识别突触体制剂以及离散脑区域和神经元凋亡诱导模型中的caspase靶点。该分析不仅可以确定突触中半胱天冬酶的蛋白质底物，还可以揭示蛋白质水解裂解的精确位点，为裂解事件的功能提供直接的分子洞察力。该项目获得的信息不仅将刺激对半胱天冬酶切割突触靶点的功能作用的进一步研究，而且还将为人类神经系统疾病（包括创伤、中风和神经退行性疾病）提供直接的候选生物标志物。