Mechanisms of Axon Pathology in ALS

ALS 轴突病理学机制

基本信息

批准号：
9927699
负责人：
SERGE E PRZEDBORSKI
金额：
$ 53.41万
依托单位：
COLUMBIA UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-09-01 至 2023-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9927699
关键词：
Adult Amyotrophic Lateral Sclerosis Anatomy Animals Automobile Driving Axon BAX gene Behavior monitoring Behavioral Cell Death Cell Membrane Proteins Clinical Cost of Illness Data Denervation Dependovirus Disease Electrophysiology (science)Enzymes Functional disorder Genetic Goals In Vitro Individual Knowledge Lead Light Lipids Longevity Mediating Modeling Molecular Morphology Motor Motor Neuron Disease Motor Neurons Mus Muscle Muscle Fibers Muscle Weakness Muscle denervation procedure Nature Nerve Neurologic Neuromuscular Junction Outcome Paralysed Pathology Pathway interactions Patients Pharmacology Phase Phenotype Post-Translational Protein Processing Preventive Preventive Intervention Process Protein Isoprenylation Proteins Proteome Quality of life Refractory Reporting Research Resistance Respiratory Muscles Role Specificity Spinal Symptoms Testing Therapeutic Therapeutic Intervention Time Transferase Transgenic Mice Transgenic Organisms Virulence Factors Withdrawal Work axon growth axonal degeneration base brain cell genetic approach improved in vitro Model in vivo innovation knock-down loss of function motor deficit mutant nerve supply neuron loss novel therapeutic intervention prenyl prenylation prevent programs protein farnesyltransferase protein geranylgeranyltransferase protein protein interaction reinnervation screening small hairpin RNA superoxide dismutase 1 therapeutic development

项目摘要

Amyotrophic lateral sclerosis (ALS) is a fatal paralytic disease characterized by neuromuscular junction (NMJ) denervation that precedes spinal motor neuron (MN) death and muscle weakness. We hypothesize that preventing denervation and stimulating reinnervation of NMJs will thwart muscle dysfunction and weakness in ALS, hence improving the patient's quality of life and, likely extending survival. Herein, we seek to demonstrate that protein prenylation, which was reported to operate as an endogenous brake on axonal growth, is a key determinant of ALS-related motor axon pathology. In support of this goal, our pilot work shows that dually silencing the prenylation enzymes, farnesyl transferase and geranylgeranyl transferase type-I, or uniquely silencing geranylgeranyl transferase type-II, mitigates NMJ denervation in the transgenic (Tg) mouse expressing mutant SOD1 (mSOD1). The rationale for this project is that, once it is known which prenylated proteins are essential for ALS-related motor axon pathology and which prenyl transferases catalyze their prenylation, new and innovative strategies can be devised for the treatment of ALS. Thus, the following three specific aims are proposed. In AIM 1, we will identify the prenyl transferase involved in motor axon pathology by silencing these enzymes individually or in combination in Tg mSOD1 mice and then, we will compare, at different time points, the number of lumbar and phrenic MNs and the NMJ innervation of ambulatory and respiratory muscles that are critical to the quality of life and lifespan, respectively. We will also demonstrate the generic nature of protein prenylation in ALS-related motor axon pathology by assessing the most effective silencing identified above in a non-SOD1 model of ALS. In AIM 2, we will ascertain the specificity of protein prenylation for motor axon pathology by monitoring behavioral, electrophysiological and anatomical parameters in Tg mSOD1 mice deficient in the pro-cell death gene Bax with and without prenylation inhibition. Since Bax deletion abrogates spinal MN death but not motor axon pathology in these mice, Tg mSOD1/Bax–/– animals will enable us to determine whether: (i) motor axon pathology and MN death are governed by distinct molecular programs and (ii) inhibition of both prenylation and Bax not only delays the onset of motor deficit but also extends lifespan. In AIM 3, we will elucidate the specific prenylated proteins that contribute to motor axon pathology by generating the MN prenylated proteome and then, use this information to perform a loss-of- function screening in an in vitro model of ALS-like axon pathology. Lastly, those silenced MN prenylated proteins that mitigate the axon phenotype in vitro will be validated in Tg mSOD1 mice using the same tests as in AIM 2. In light of the above, we expect that the successful completion of the proposed work will identify the prenylation pathway and its targets that contribute to motor axon pathology in ALS. These findings will have an important positive impact in that they will provide opportunities for preventive and therapeutic interventions and, fundamentally, advance our mechanistic understanding of ALS and related disorders.

肌萎缩侧索硬化症（ALS）是一种致命性麻痹性疾病，其特征是神经肌肉接头（NMJ）脊髓运动神经元（MN）死亡和肌肉无力之前的去神经支配。预防 NMJ 的去神经支配并刺激 NMJ 的神经支配再支配将阻止肌肉功能障碍和肌无力 ALS，从而改善患者的生活质量，并可能延长生存期。据报道，蛋白质异戊二烯化是轴突生长的一个关键因素。 ALS 相关运动轴突病理学的决定因素为了支持这一目标，我们的试点工作表明了这一点。沉默异戊二烯化酶、法尼基转移酶和香叶基香叶基转移酶 I 型，或独特地沉默香叶基香叶基转移酶 II 型，减轻转基因 (Tg) 小鼠的 NMJ 去神经支配表达突变体 SOD1 (mSOD1) 该项目的基本原理是，一旦知道哪个异戊二烯化。蛋白质对于 ALS 相关运动轴突病理学至关重要，并且异戊烯基转移酶催化其异戊二烯化可以设计出新的创新策略来治疗 ALS，因此，有以下三种策略。在 AIM 1 中，我们将确定参与运动轴突病理学的异戊二烯基转移酶。通过在 Tg mSOD1 小鼠中单独或组合沉默这些酶，然后我们将进行比较不同时间点、腰椎和膈肌 MN 的数量以及动态和运动的 NMJ 神经支配我们还将展示分别对生活质量和寿命至关重要的呼吸肌。通过评估最有效的方法，了解 ALS 相关运动轴突病理学中蛋白质异戊二烯化的一般性质在 AIM 2 中，我们将确定上述 ALS 非 SOD1 模型中蛋白质的特异性。通过监测行为、电生理和解剖参数来进行运动轴突病理学的异戊二烯化在具有或不具有自 Bax 抑制的亲细胞死亡基因 Bax 的 Tg mSOD1 小鼠中。缺失消除了脊髓 MN 死亡，但没有消除这些小鼠的运动轴突病理学，Tg mSOD1/Bax–/– 动物将使我们能够确定：（i）运动轴突病理学和 MN 死亡是否由不同的分子控制 (ii) 抑制异戊二烯化和 Bax 不仅可以延迟运动缺陷的发生，还可以在 AIM 3 中，我们将阐明有助于运动轴突的特定异戊烯化蛋白。通过生成 MN 异戊二烯化蛋白质组来进行病理学分析，然后使用此信息执行丢失- 在类似 ALS 的轴突病理学体外模型中进行功能筛选最后，那些沉默的 MN 异戊二烯化。体外减轻轴突表型的蛋白质将在 Tg mSOD1 小鼠中使用与在 AIM 2 中。鉴于上述情况，我们预计拟议工作的成功完成将确定异戊二烯化途径及其有助于 ALS 运动轴突病理学的靶标这些发现将具有重要意义。重要的积极影响，因为它们将为预防和治疗干预提供机会从根本上推进我们对 ALS 及相关疾病的机制理解。