Defining the impact of Frataxin point mutations on Friedreich's ataxia pathogenesis

弗里德赖希共济失调（FRDA）是一种常染色体隐性遗传性神经退行性疾病， 线粒体蛋白共济失调蛋白（FXN）。Frataxin翻译为210个氨基酸（aa）前体（FXN-P） 它被输入到线粒体基质中，在那里它经历连续的切割步骤，产生168个氨基酸， 中间体（FXN-I）和129 aa的成熟同种型（FXN-M）。Frataxin参与铁硫簇（ISC） 线粒体中的生物合成，许多明显的FRDA表型是由ISC-1的活性缺陷引起的。 含有酶。目前，没有治愈这种使人衰弱的疾病。大多数FRDA患者 FXN基因内含子1中GAA三联体重复序列的大扩增是纯合的，而 的患者是复合杂合子，其中一个FXN等位基因中的GAA重复序列扩增， 或无意义突变。纯合子和复合杂合子突变基因型均导致 与健康对照组相比，FXN-M蛋白水平降低。最普遍的错义突变 将第130位的甘氨酸变为缬氨酸（G130 V）。FRDA G130 V患者表现出与 携带纯合子GAA扩增的患者，包括下肢痉挛而非共济失调， 感觉反应、言语和上肢功能保留以及疾病进展减慢。巧合的是， 在G130 V患者样品中可检测到的FXN-M蛋白显著少于携带两种FXN-M蛋白的患者样品。 扩展等位基因我们的初步数据显示，FXN的正常线粒体成熟过程 蛋白质被G130 V突变扰乱，表明中间异构体的功能重要性 （G130V-I）。我们假设G130 V突变损害FXN线粒体成熟过程和/或 使成熟的同种型不稳定。未处理的FXN-G130 V-I同种型具有功能性并部分补偿 对于FXN-M的大幅减少，从而减缓疾病进展并有助于不同的免疫反应。 FRDA G130 V患者的症状。为了解决这些假设，我们将使用新的细胞和小鼠模型 FRDA G130V首先，我们将定义FXN-G130 V-I同种型的结构和功能特性以测试 该突变是否赋予功能改变，导致FRDA的非典型临床表现 G130 V患者。随后，我们将确定控制稳态水平和成熟的机制 在iPSC衍生的皮层和感觉神经元中处理FXN-G130 V。最后，使用FRDA患者源性 神经元模型以及我们的新型Fxn G127 V小鼠模型，我们将定义潜在的分子机制， FRDA G130 V患者的独特临床表现。拟议研究的结果将具有广泛的 对所有FRDA患者治疗开发的影响。

Friedreich’s ataxia (FRDA) is an autosomal recessive neurodegenerative disease caused by reduced expression of the mitochondrial protein frataxin (FXN). Frataxin is translated as a 210 amino acid (aa) precursor (FXN-P) that is imported into the mitochondrial matrix where it undergoes sequential cleavage steps, producing a 168 aa intermediate (FXN-I) and the mature isoform of 129 aa (FXN-M). Frataxin participates in iron-sulfur cluster (ISC) biosynthesis in the mitochondria, and many of the overt FRDA phenotypes result from deficient activity of ISC- containing enzymes. Currently, there is no cure for this debilitating disease. Most FRDA patients are homozygous for large expansions of GAA triplet repeat sequences in intron 1 of the FXN gene, while a subset of patients are compound heterozygotes with an expanded GAA repeat tract in one FXN allele and a missense or nonsense mutation in the other. Homozygous and compound heterozygous mutant genotypes both result in reduced levels of FXN-M protein when compared with healthy controls. The most prevalent missense mutation changes a glycine to valine at position 130 (G130V). FRDA G130V patients exhibit different clinical features than patients harboring homozygous GAA expansions, including lower limb spasticity rather than ataxia, preserved sensory responses, spared speech and upper limb functions, and slower disease progression. Paradoxically, substantially less FXN-M protein is detectable in G130V patient samples than in patient samples harboring two expanded alleles. Our preliminary data revealed that normal mitochondrial maturation processing of the FXN protein is perturbed by the G130V mutation, suggesting functional importance of an intermediate isoform (G130V-I). We hypothesize that the G130V mutation impairs FXN mitochondrial maturation processing and/or destabilizes the mature isoform. The unprocessed FXN-G130V-I isoform is functional and partially compensates for the substantial reduction of FXN-M, thus slowing disease progression and contributing to the distinct symptoms of FRDA G130V patients. To address these hypotheses, we will use novel cellular and mouse models of FRDA G130V. First, we will define the structural and functional properties of the FXN-G130V-I isoform to test whether this mutation confers a change of function that contributes to the atypical clinical presentation of FRDA G130V patients. Subsequently, we will determine mechanisms governing steady state levels and maturation processing of FXN-G130V in iPSC-derived cortical and sensory neurons. Finally, using FRDA patient-derived neuronal models as well as our novel Fxn G127V mouse model, we will define molecular mechanisms underlying the unique clinical presentation of FRDA G130V patients. Results of the proposed studies will have a broad impact on therapy development for all FRDA patients.