An in vivo approach to understanding mutant PFN1 toxicity on motor neurons

项目摘要 ALS，也称为Lou Gehrig病，是一种无法治愈的神经退行性疾病， 运动神经元的丧失导致瘫痪并最终死亡。为了了解这种疾病 机制和开发治疗方法，哺乳动物模型，表型人类疾病是 不可或缺二十多年来，表达突变SOD 1基因的转基因动物一直受到关注。 这是唯一一个能忠实地代表人类疾病并发挥重要作用的模型 在推进我们对ALS的理解和促进治疗发展方面。但由于 由于缺乏来自其他ALS致病基因的动物模型，因此很难进行验证和推广 突变SOD 1模型的机制和治疗发现。因此，我们不 了解SOD1模型的机制发现是否对不同的突变体是共同的 基因或其仅特异于SOD1突变。这阻碍了我们对ALS的理解， 治疗发展因此，模拟ALS的另外的哺乳动物模型的构建 在人类疾病的过程中是我们对抗ALS的关键。然而，在开发新的 具有导致运动神经元丧失的进行性ALS表型的哺乳动物模型，临床 瘫痪和死亡是很困难的为了解决这个问题，我们构建了一个转基因的 通过表达新近发现的ALS基因突变profilin1基因（PFN1C71G）建立小鼠模型。我们 显示突变体而非野生型（PFN1WT）基因表达引起迟发性运动 功能障碍表型，随后发展为瘫痪和死亡。而且 突变小鼠的运动神经元变性不断发展并失去大多数 运动神经元的功能。这些结果表明突变型PFN1导致ALS 通过毒性的增加，建立一个渐进的ALS疾病模型， 人类疾病这些小鼠为研究疾病机制提供了新的体内系统， ALS的治疗方法该建议利用这种新的模式，并寻求机制 对运动神经元退化的见解目的1将区分两种机制的突变 PFN1毒性，突变蛋白获得新的毒性特性与显性负抑制 正常PFN1等位基因目的2将探讨潜在的细胞决定因素的疾病 发病率和疾病进展率。目的3探讨内质网损伤在突变型 PFN1诱导的运动神经元变性。我们将把这些目标的结果与 从突变的SOD1模型的发现，以确定共同的或基因特异性机制。由这些 了解后，可以实现更好的治疗方法。

PROJECT SUMMARY ALS, also known as Lou Gehrig’s disease, is an incurable neurodegenerative disease caused by the loss of motor neurons leading to paralysis and eventually death. To understand the disease mechanism and develop therapeutics, mammalian models that phenocopy human disease are indispensable. For more than two decades, transgenic animals expressing mutant SOD1 gene were the only model available that faithfully represented the human disease and played an essential role in advancing our understandings of ALS and enabling therapeutic development. However, due to the lack of animal models from other ALS-causal genes, it has been difficult to verify and generalize the mechanistic and therapeutic findings from the mutant SOD1 models. Consequently, we do not know whether the mechanistic findings from the SOD1 models are common to different mutant genes or it is specific for SOD1 mutations alone. This has hampered our understanding of ALS and therapeutic development. Thus, construction of additional mammalian models that mimic ALS disease process in human is crucial in our fight against ALS. However, efforts in developing new mammalian models with progressive ALS phenotypes leading to motor neuron loss, clinical paralysis and death has proven difficult. To solve this problem, we have constructed a transgenic mouse model by expressing mutant profilin1 gene (PFN1C71G), a recently discovered ALS gene. We show that expression of the mutant, but not the wild type (PFN1WT) gene, caused a late onset motor dysfunction phenotype that subsequently progressed to paralysis and death. Furthermore, the mutant mice developed a relentless progression of motor neuron degeneration and lose a majority of their motor neurons at the end stage. These results demonstrate that mutant PFN1 causes ALS by a gain of toxicity and establish a progressive ALS disease model that closely phenocopy the human disease. These mice provide a new in vivo system for study of disease mechanisms and therapeutics for ALS. This proposal take advantage of this new model and seeks mechanistic insights on motor neuron degeneration. Aim 1 will differentiate between two mechanisms of mutant PFN1 toxicity, a gain of novel toxic property by the mutant protein vs. a dominant-negative inhibition of the normal PFN1 allele. Aim 2 will explore the underlying cellular determinants for the disease onset and the rate of disease progression. Aim 3 will investigate the role of ER damage in mutant PFN1-induced motor neuron degeneration. We will compare the findings from these aims with the findings from mutant SOD1 models to determine common or gene-specific mechanisms. By these understanding, better therapeutic approaches may be achieved.