Mechanisms of Axon Pathology in ALS

ALS 轴突病理学机制

基本信息

批准号：
10403431
负责人：
SERGE E PRZEDBORSKI
金额：
$ 51.81万
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 2024-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10403431
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 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 amyotrophic lateral sclerosis therapy 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 preventive intervention 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.

肌萎缩性外侧硬化症（ALS）是一种致命的麻痹性疾病，其特征是神经肌肉结（NMJ）在脊柱运动神经元（MN）死亡和肌肉无力之前进行的改造。我们假设这一点防止神经支配和刺激NMJ的重新连接将阻止肌肉功能障碍和无力 ALS，因此改善了患者的生活质量，并可能延长生存。在这里，我们试图证明据报道，该蛋白介载是在轴突生长时作为内源制动的，是钥匙与ALS相关的运动轴突病理学的决定因素。为了支持这个目标，我们的飞行员工作表明沉默原型酶，Farnesyl转移和黄烷基转移酶I型或独特的沉默的黄烷基转移酶II型，减轻转基因（TG）小鼠中的NMJ神经化。表达突变体SOD1（MSOD1）。该项目的理由是，一旦知道了蛋白质对于与ALS相关的运动轴突病理学至关重要，并且蛋白质可促催化其催化其可以设计用于治疗ALS的新策略，新的和创新的策略。那，以下三个提出了具体目标。在AIM 1中，我们将确定与运动轴突病理相关的前转移通过单独或组合TG MSOD1小鼠中的这些酶沉默，然后我们将在不同的时间点，腰部和Phrenic MN的数量以及卧床的NMJ神经支配分别对生活质量和寿命至关重要的呼吸道肌肉。我们还将证明通过评估最有效上面在非SOD1模型的ALS模型中确定的沉默。在AIM 2中，我们将确定蛋白质的特异性通过监测行为，电生理和解剖参数，用于运动轴突病理的原始化在TG MSOD1小鼠中，缺乏促核细胞死亡基因BAX的小鼠有或不具有前介苯抑制。自BAX以来删除这些小鼠中的脊柱MN死亡，但不能消除运动轴突病理学，TG MSOD1/BAX - / - 将使我们能够确定：（i）运动轴突病理学和MN死亡由不同的分子控制程序和（ii）抑制前化和BAX的抑制不仅延迟了运动不足的发作，而且还延迟延长寿命。在AIM 3中，我们将阐明有助于运动轴突的特定前蛋白通过产生Mn的蛋白质组来病理学，然后使用此信息执行损失在ALS样轴突病理学的体外模型中进行功能筛选。最后，那些沉默的Mn序列化在TG MSOD1小鼠中，使用与相同的测试与相同的测试验证了减轻轴突表型的蛋白在AIM 2中。鉴于上述，我们期望拟议的工作的成功完成将确定促进途径及其靶标有助于ALS中的运动轴突病理。这些发现将有一个重要的积极影响，因为它们将为预防和治疗干预提供机会而且，从根本上讲，我们对ALS和相关疾病的机械理解提高了。