The role of glycyl-tRNA synthetase in axonal health and disease

 描述(申请人提供)：我作为一名内科科学家的职业生涯的一个主要目标是了解遗传性神经退行性疾病的机制，最终目的是开发治疗方法。Charcot-Marie-Tooth(CMT)病是最常见的遗传性周围神经病，全世界每2500人中就有一人受到影响。到目前为止，已有6个氨酰-tRNA合成酶(ARS)基因与CMT病的轴突病理有关。在常染色体显性遗传性轴索性CMT 2型(CMT2)患者中，已发现5个ARS基因座的21个突变。到目前为止，大多数与疾病相关的ARS等位基因要么干扰了酶的催化活性，要么错误地定位了神经元中的酶，从而导致功能丧失。这表明ARS功能受损是CMT2病的一个重要方面。有趣的是，大多数与CMT相关的ARS突变都是错义突变；移码和无义突变在显性遗传性CMT2疾病中尚未被发现。这些观察导致了我们的假设，即显性-负性效应可能是轴突表型的原因；然而，显性-负性机制尚未得到评估。内源性甘氨酰-tRNA合成酶(GARS)在细胞中形成离散的点状结构，包括在人类周围神经的轴突中。在与CMT病相关的12个GARS突变中，有7个在体外阻止了点状突起的形成，表明这些点状突起可能对轴突的健康和维持很重要。然而，斑点的组成和功能尚不清楚。我们假设GARS与这些斑点中的翻译机制的组件相关联，以促进轴突内高效的蛋白质翻译。为了解决这些关键问题，我们将：(1)直接评估GARS突变在体内的显性负面影响；以及(2)表征GARS斑点在轴突中的功能。这些实验将提高我们对GARS相关CMT疾病的理解，并将为治疗开发提供信息。重要的是，这个项目将让我发展必要的技能，为专注于了解神经退行性疾病机制的研究生涯提供必要的技能。

 DESCRIPTION (provided by applicant): A major goal of my career as a physician-scientist is to understand the mechanisms of inherited neurodegenerative diseases, with the ultimate aim of developing therapies. Charcot-Marie-Tooth (CMT) disease is the most common inherited peripheral neuropathy, affecting one in 2,500 individuals worldwide. To date, six aminoacyl-tRNA synthetase (ARS) genes have been implicated in CMT disease with an axonal pathology. Twenty-one mutations at five ARS loci have been identified in patients with autosomal dominant, axonal CMT type 2 (CMT2) disease. To date, the majority of disease-associated ARS alleles cause a loss of function by either interfering with the catalytic activity or mislocalizingthe enzyme in neurons. This indicates that impaired ARS function is an important aspect of CMT2 disease. Interestingly, the majority of CMT- associated ARS mutations are missense changes; frameshift and nonsense mutations have not been identified in dominantly inherited CMT2 disease. These observations led to our hypothesis that a dominant-negative effect may be responsible for the axonal phenotype; however, a dominant-negative mechanism has not been evaluated. Endogenous glycyl-tRNA synthetase (GARS) forms discrete puncta in cells, including in the axons of human peripheral nerves. Seven of the 12 GARS mutations associated with CMT disease prevent the formation of puncta in vitro, indicating that these puncta may be important for the health and maintenance of axons. However, the composition and function of the puncta are unknown. We hypothesize that GARS associates with components of the translational machinery in these puncta to promote efficient protein translation within axons. To address these critical issues, we will: (1) directly evaluate GARS mutations for a dominant-negative effect in vivo; and (2) characterize the function of GARS puncta in axons. These experiments will improve our understanding of GARS-related CMT disease and will inform therapeutic development. Importantly, this project will allow me to develop the skill set necessary for a research career focused on understanding the mechanisms of neurodegenerative diseases.