Exploring Apoptosome-Independent Mechanisms for Casp9 activation in Axon Pruning

探索轴突修剪中 Casp9 激活的独立凋亡体机制

• 批准号: 10288453
• 负责人: Mohanish P Deshmukh
• 金额: $ 42.76万
• 依托单位: UNIV OF NORTH CAROLINA CHAPEL HILL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10288453
• 关键词: Active Sites; Adult; Alzheimer' s Disease; Apoptosis; Axon; BIRC4 gene; Binding; Biological; CASP3 gene; CASP9 gene; Caspase; Cell Death; Cells; Cessation of life; Cleaved cell; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Dependence; Development; Dimerization; Ensure; FHL2 gene; Goals; Huntington Disease; Impairment; Interphase Cell; Knock-out; Mediating; Microfluidics; Mitochondria; Modeling; Molecular; Mus; Mutation; Nerve Growth Factors; Nervous system structure; Neurodegenerative Disorders; Neurons; Outer Mitochondrial Membrane; Parkinson Disease; Pathway interactions; Peripheral; Physiological; Proteins; Reporting; Risk; Signal Transduction; Site; apoptotic protease-activating factor 1; aposome; axonal degeneration; base; clinically relevant; cytochrome c; deprivation; inhibitor/antagonist; knock-down; mimetics; mutant; neuron apoptosis; neuronal cell body; novel; receptor; small hairpin RNA; virtual

Project Summary/Abstract Neurons have the capability to activate pathways that cause degeneration of either the entire cell by apoptosis or only the axons. Physiological axon-specific degeneration, also known as pruning, is important as it allows neurons to remove excessive or misguided axon branches and permit plasticity in neuronal connections. However, exactly how a neuron can activate and compartmentalize this pathway to degenerate its axon without putting the rest of the cell at risk is unclear. This is particularly interesting since recent studies identified Bax and caspases, key proteins in the apoptosis pathway, to also regulate axon pruning We are using a microfluidic chamber-based model of sympathetic neurons where deprivation of nerve growth factor (NGF) can induce either apoptosis (when NGF is deprived from both soma and axon compartments) or axon pruning (when NGF is deprived from only the axon compartment). Using this model, we identified substantial overlap but also distinct differences between the apoptosis and axon pruning pathways. Specifically, we found that while caspase-9 (Casp9) is required for both pathways, Casp9 activation is dependent of Apaf-1 during apoptosis but, surprisingly, independent of Apaf-1 during axon pruning. These results were unexpected and point to a novel mechanism of Casp9 activation during axon pruning. In this proposal, our goals are to examine specific aspects of how Casp9 is activated during axon pruning. In Aim 1, we will focus on Bax function during axon pruning. Since Apaf-1 is not needed for axon pruning, we propose that the essential function of Bax during axon pruning is not the release of cyt c (which activates Apaf- 1 in the context of apoptosis), but instead is the release of the Smac, an inhibitor of XIAP, from mitochondria. In Aim 2, we will examine how Casp9 is activated during pruning. Our focus is on identifying key features of Casp9 function that are important for axon pruning. Additionally, we will examine whether components of a dependosome-like complex are important for activating Casp9 during axon pruning. These studies will help uncover critical aspects of how neurons utilize many of the same components for apoptosis and axon pruning yet with distinct differences.

项目总结/摘要 神经元有能力激活导致整个细胞退化的途径， 凋亡或仅轴突。生理性轴突特异性变性，也称为修剪，是重要的， 它允许神经元去除过多的或错误的轴突分支，并允许神经元的可塑性， 连接.然而，神经元究竟如何激活和划分这一通路，使其退化， 轴突而不将细胞的其他部分置于危险之中还不清楚。这是特别有趣的，因为最近的研究 确定了凋亡途径中的关键蛋白Bax和半胱天冬酶，也调节轴突修剪 我们正在使用一种基于微流体室的交感神经元模型， 当索马和轴突都缺乏NGF时， 轴突修剪（当仅从轴突隔室剥夺NGF时）。利用这个模型， 我们确定了大量的重叠，但也有明显的差异，细胞凋亡和轴突修剪 途径。具体地说，我们发现，虽然caspase-9（Casp 9）是两种途径所必需的，但Casp 9的活化 在凋亡过程中依赖于Apaf-1，但令人惊讶的是，在轴突修剪过程中不依赖于Apaf-1。这些 结果是出乎意料的，并指出了轴突修剪过程中Casp 9活化的新机制。 在这项提案中，我们的目标是研究在轴突修剪过程中Casp 9如何被激活的具体方面。 在目标1中，我们将专注于轴突修剪过程中的Bax功能。由于轴突修剪不需要Apaf-1，我们 提出Bax在轴突修剪过程中的基本功能不是释放cyt c（其激活Apaf）， 1在细胞凋亡的情况下），而是Smac，XIAP的抑制剂，从线粒体的释放。 在目标2中，我们将研究在修剪过程中如何激活Casp 9。我们的重点是确定关键特征， 对轴突修剪很重要的Casp 9功能。此外，我们将研究是否组件的 依赖体样复合物对于轴突修剪期间激活Casp 9是重要的。这些研究将有助于 揭示神经元如何利用许多相同的成分进行细胞凋亡和轴突修剪的关键方面 但有明显的不同。