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

确定 Frataxin 点突变对 Friedreich 共济失调发病机制的影响

• 批准号: 10563061
• 负责人: Marek Napierala
• 金额: $ 41万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-01 至 2026-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10563061
• 关键词: Address; Affect; Afferent Neurons; Alleles; Amino Acids; Anabolism; Ataxia; Axonal Transport; Binding; Biochemical; Biophysics; Cell model; Cell physiology; Cells; Characteristics; Clinical; Complex; Data; Disease; Disease Progression; Electron Microscopy; Enzymes; Exhibits; Friedreich Ataxia; Genes; Genotype; Glycine; Hereditary Spastic Paraplegia; Heterozygote; Impairment; In Vitro; Introns; Iron; Knowledge; Link; Lower Extremity; Mediating; Missense Mutation; Mitochondria; Mitochondrial Matrix; Mitochondrial Proteins; Modeling; Molecular; Molecular Chaperones; Molecular Sieve Chromatography; Mutation; NMR Spectroscopy; Neurodegenerative Disorders; Neurons; Nonsense Mutation; Pathogenesis; Pathogenicity; Pathologic; Pathway interactions; Patients; Peptide Hydrolases; Phenotype; Point Mutation; Positioning Attribute; Production; Property; Protein Isoforms; Proteins; Reagent; Respiration; Sampling; Sensory; Severities; Speech; Sulfur; Symptoms; Testing; Therapeutic; Translating; Trinucleotide Repeats; Upper Extremity; Valine; Work; axonal degeneration; base; design; experience; experimental study; frataxin; induced pluripotent stem cell; lipid metabolism; mouse model; multiple myeloma M Protein; mutant; novel; patient subsets; preservation; response; spasticity; therapy development

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.

弗里德赖希共济失调 (FRDA) 是一种由表达减少引起的常染色体隐性神经退行性疾病 线粒体蛋白 frataxin (FXN)。 Frataxin 被翻译为 210 个氨基酸 (aa) 前体 (FXN-P) 被导入线粒体基质中，在那里经历连续的裂解步骤，产生 168 个氨基酸 中间体 (FXN-I) 和 129 个氨基酸的成熟亚型 (FXN-M)。 Frataxin 参与铁硫簇 (ISC) 线粒体中的生物合成，许多明显的 FRDA 表型是由于 ISC- 活性缺陷造成的 含有酶。目前，这种使人衰弱的疾病无法治愈。大多数 FRDA 患者是 FXN 基因内含子 1 中 GAA 三联体重复序列的大量扩展是纯合的，而一个子集 的患者是复合杂合子，在一个 FXN 等位基因中具有扩展的 GAA 重复序列和错义 或对方的无义突变。纯合子和复合杂合子突变基因型都会导致 与健康对照相比，FXN-M 蛋白水平降低。最常见的错义突变 将 130 位的甘氨酸更改为缬氨酸 (G130V)。 FRDA G130V 患者表现出与其他患者不同的临床特征 携带纯合 GAA 扩展的患者，包括下肢痉挛而不是共济失调，保留 感觉反应，言语和上肢功能不受损害，并且疾病进展较慢。矛盾的是， 在 G130V 患者样本中检测到的 FXN-M 蛋白比在含有两种蛋白的患者样本中检测到的 FXN-M 蛋白要少得多 扩展的等位基因。我们的初步数据显示，FXN 的正常线粒体成熟过程 蛋白质受到 G130V 突变的干扰，表明中间亚型的功能重要性 (G130V-I)。我们假设 G130V 突变损害 FXN 线粒体成熟处理和/或 破坏成熟亚型的稳定性。未处理的 FXN-G130V-I 同工型具有功能性并部分补偿 FXN-M 的大幅减少，从而减缓疾病进展并有助于明显的 FRDA G130V 患者的症状。为了解决这些假设，我们将使用新型细胞和小鼠模型 FRDA G130V。首先，我们将定义 FXN-G130V-I 同工型的结构和功能特性进行测试 这种突变是否会导致功能改变，从而导致 FRDA 的非典型临床表现 G130V患者。随后，我们将确定控制稳态水平和成熟度的机制 FXN-G130V 在 iPSC 衍生的皮质和感觉神经元中的处理。最后，使用 FRDA 患者衍生的 神经元模型以及我们的新型 Fxn G127V 小鼠模型，我们将定义潜在的分子机制 FRDA G130V 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。