Friedreich Ataxia (FA) is caused by GAA repeat expansions in the first intron of FXN, the gene encoding frataxin, which results in decreased gene expression. Thanks to the high degree of frataxin conservation, the Drosophila melanogaster fruitfly appears as an adequate animal model to study this disease and to evaluate therapeutic interventions. Here, we generated a Drosophila model of FA with CRISPR/Cas9 insertion of approximately 200 GAA in the intron of the fly frataxin gene fh. These flies exhibit a developmental delay and lethality associated with decreased frataxin expression. We were able to by-pass preadult lethality using genetic tools to overexpress frataxin only during the developmental period. These frataxin-deficient adults are short-lived and present strong locomotor defects. RNA-Seq analysis identified deregulation of genes involved in amino-acid metabolism and transcriptomic signatures of oxidative stress. In particular, we observed a progressive increase of Tspo expression, fully rescued by adult frataxin expression. Thus, Tspo expression constitutes a molecular marker of the disease progression in our fly model and might be of interest in other animal models or in patients. Finally, in a candidate drug screening, we observed that N-acetyl cysteine improved the survival, locomotor function, resistance to oxidative stress and aconitase activity of frataxin-deficient flies. Therefore, our model provides the opportunity to elucidate in vivo the protective mechanisms of this molecule of therapeutic potential. This study also highlights the strength of the CRISPR/Cas9 technology to introduce human mutations in endogenous orthologous genes, leading to Drosophila models of human diseases with improved physiological relevance.

中文翻译：

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在 frataxin 基因中插入 GAA 重复序列的 CRISPR/Cas9 的 Friedreich Ataxia 果蝇模型揭示了 N-乙酰半胱氨酸的体内保护。

Friedreich Ataxia (FA) 是由FXN的第一个内含子中的 GAA 重复扩增引起的，FXN是编码 frataxin 的基因，导致基因表达下降。由于 frataxin 的高度保护，黑腹果蝇果蝇似乎是研究这种疾病和评估治疗干预的合适动物模型。在这里，我们生成了 FA 的果蝇模型，在果蝇 frataxin 基因fh的内含子中插入了大约 200 GAA 的 CRISPR/Cas9. 这些果蝇表现出与 frataxin 表达降低相关的发育延迟和致死率。我们能够使用遗传工具仅在发育期间过度表达 frataxin 来绕过成年前的致死率。这些缺乏 frataxin 的成年人寿命很短，并且存在严重的运动缺陷。RNA-Seq 分析确定了参与氨基酸代谢和氧化应激转录组特征的基因的失调。特别是，我们观察到Tspo表达的逐渐增加，完全被成人 frataxin 表达所拯救。因此，Tspo表达构成了我们果蝇模型中疾病进展的分子标记，并且可能对其他动物模型或患者感兴趣。最后，在候选药物筛选中，我们观察到 N-乙酰半胱氨酸提高了 frataxin 缺陷果蝇的存活率、运动功能、对氧化应激的抵抗力和乌头酸酶活性。因此，我们的模型提供了在体内阐明这种具有治疗潜力的分子的保护机制的机会。这项研究还突出了 CRISPR/Cas9 技术在内源直系同源基因中引入人类突变的优势，从而导致人类疾病的果蝇模型具有更高的生理相关性。