Regulation of mitochondrial dehdyrogenases and neuronal NADH levels via interaction of NIPSNAP1 and APP intracellular domain

通过 NIPSNAP1 和 APP 胞内域相互作用调节线粒体脱氢酶和神经元 NADH 水平

基本信息

批准号：
9318084
负责人：
RAMIN HOMAYOUNI
金额：
$ 21.07万
依托单位：
UNIVERSITY OF MEMPHIS
依托单位国家：
美国
项目类别：
财政年份：
2017
资助国家：
美国
起止时间：
2017-04-01 至 2019-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9318084
关键词：
Affect Affinity Aging Alzheimer&apos s Disease Amyloid beta-Protein Amyloid beta-Protein Precursor Amyotrophic Lateral Sclerosis Apoptosis Binding Biochemical Biological Assay Brain Cell physiology Cells Complex Cytoplasmic Tail Data Enzymes Equilibrium Genes Genetic Genetically Engineered Mouse Goals Hippocampus (Brain)Huntington Disease In Vitro Inbred Strains Mice Keto Acids Lead Ligands Link Liver Maintenance Metabolism Mitochondria Mitochondrial Proteins Modeling Molecular Mus Mutation NADH NADP Natural regeneration Nerve Degeneration Neurodegenerative Disorders Neurons Nucleotides Oxidoreductase Parkinson Disease Pathogenesis Pathologic Peptides Phosphoric Monoester Hydrolases Physiological Play Polyacrylamide Gel Electrophoresis Preparation Presenile Alzheimer Dementia Production Proteins Proteomics Reactive Oxygen Species Recombinants Reducing Agents Regulation Reporting Role S-nitro-N-acetylpenicillamine Survival Rate System Testing Time Tissues Western Blotting Work age related base enzyme activity excitotoxicity experimental study genetic analysis insight metabolomics mitochondrial dysfunction mouse model neuronal survival neurotoxicity novel presenilin-1 presenilin-2 protein structure pyruvate dehydrogenase screening therapeutic development virtual

Affect Affinity Aging Alzheimer&apos s Disease Amyloid beta-Protein Amyloid beta-Protein Precursor Amyotrophic Lateral Sclerosis Apoptosis Binding Biochemical Biological Assay Brain Cell physiology Cells Complex Cytoplasmic Tail Data Enzymes Equilibrium Genes Genetic Genetically Engineered Mouse Goals Hippocampus (Brain)Huntington Disease In Vitro Inbred Strains Mice Keto Acids Lead Ligands Link Liver Maintenance Metabolism Mitochondria Mitochondrial Proteins Modeling Molecular Mus Mutation NADH NADP Natural regeneration Nerve Degeneration Neurodegenerative Disorders Neurons Nucleotides Oxidoreductase Parkinson Disease Pathogenesis Pathologic Peptides Phosphoric Monoester Hydrolases Physiological Play Polyacrylamide Gel Electrophoresis Preparation Presenile Alzheimer Dementia Production Proteins Proteomics Reactive Oxygen Species Recombinants Reducing Agents Regulation Reporting Role S-nitro-N-acetylpenicillamine Survival Rate System Testing Time Tissues Western Blotting Work age related base enzyme activity excitotoxicity experimental study genetic analysis insight metabolomics mitochondrial dysfunction mouse model neuronal survival neurotoxicity novel presenilin-1 presenilin-2 protein structure pyruvate dehydrogenase screening therapeutic development virtual

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项目摘要

Abstract Mitochondrial dysfunction is associated with ageing as well as a number of age-related neurodegenerative diseases including Alzheimer’s Disease (AD). Early onset AD has been linked to mutations in amyloid precursor protein (APP) and presenilins 1 and 2 (PS1 and PS2), which result in abnormal cleavage of APP and release of toxic amyloid beta (Apeptides. Accumulating evidence suggests that APP intracellular domain (AICD) may also contribute to pathogenesis of AD. To gain insights into the normal and pathological roles of AICD, we previously used a biochemical affinity proteomic strategy and found that AICD directly interacts with a novel mitochondrial protein, Nipsnap1 (4-nitrophenyl phosphatase domain and non-neuronal SNAP-25 like protein homolog1). Although Nipsnap1 is evolutionarily conserved, very little is known about its function. Our long-term goal is to investigate the molecular and cellular function of Nipsnap1 and to determine its role in neurodegeneration. Toward this end, we generated a mouse with a targeted disruption of the Nipsnap1 gene. Disruption of Nipsnap1 expression profoundly affects intermediate metabolism and significantly increased apoptosis and neurodegeneration in the brain. Protein structure modeling and virtual ligand screening suggested that Nipsnap1 may bind to NADH and NADPH. Using in vitro biochemical assays, we found for the first time that Nipsnap1 directly binds to both NADH and NADPH. Moreover, we found significantly lower NAD+/NADH ratios in Nipsnap1 deficient brain. The balance between NAD+ and NADH is critical for production of ATP, maintenance of mitochondrial potential and regeneration of reducing agents within cells to counteract reactive oxygen radicals. Based on these preliminary results, we hypothesize that Nipsnap1 plays an important role in neuronal survival by modulating dehydrogenase activities and NAD(P)H levels. In this project, we will use biochemical approaches and primary neuronal cultures derived from WT and Nipsnap1 deficient mice to determine if: 1) Nipsnap1 interacts with and regulates multiple dehydrogenases in the mitochondria; 2) AICD interaction with Nipsnap1 affects dehydrogenase activity and neuronal NAD+/NADH levels. Our work will provide insights into the molecular function of Nipsnap1 and possibly a new mechanism by which AICD produces neurotoxicity.

抽象的线粒体功能障碍与衰老以及许多与年龄相关的神经退行性有关包括阿尔茨海默氏病（AD）在内的疾病。早期发作广告与淀粉样蛋白突变有关前体蛋白（APP）和Presenilins 1和2（PS1和PS2），导致APP的异常裂解和释放有毒淀粉样ββ（A肽。积累证据表明APP内域（AICD）也可能有助于AD的发病机理。为了了解对正常和病理作用的见解 AICD，我们以前使用了生化亲和力蛋白质组学策略，发现AICD直接与一种新型的线粒体蛋白NIPSNAP1（4-硝基苯基磷酸酶结构域和非神经元SNAP-25 蛋白质同源性1）。尽管NIPSNAP1在进化上是保守的，但对其功能几乎不知情。我们的长期目标是研究NIPSNAP1的分子和细胞功能，并确定其在神经变性。为此，我们生成了一个靶向破坏NIPSNAP1基因的小鼠。 NIPSNAP1表达的破坏深刻影响中间代谢，并显着增加大脑中的凋亡和神经退行性。蛋白质结构建模和虚拟配体筛选建议NIPSNAP1可能与NADH和NADPH结合。使用体外生化测定，我们发现 NIPSNAP1首次直接与NADH和NADPH结合。此外，我们发现明显较低 NAD+/NADH比率NIPSNAP1默认大脑。 NAD+和NADH之间的平衡对于产生ATP，维持线粒体潜力和细胞内还原剂的再生抵消活性氧自由基。基于这些初步结果，我们假设NIPSNAP1播放通过调节脱氢酶活性和NAD（P）H水平，在神经元存活中的重要作用。在这个项目，我们将使用源自WT和NIPSNAP1的生化方法和主要的神经元培养物不足的小鼠确定是否：1）NIPSNAP1与中的多种脱氢酶相互作用并调节线粒体； 2）AICD与NIPSNAP1的相互作用会影响脱氢酶活性和神经元NAD+/NADH 水平。我们的工作将提供有关NIPSNAP1分子功能的见解，并可能有一种新的机制 AICD产生神经毒性。