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Rescue of Fragile X Syndrome Neurons by DNA Methylation Editing of the FMR1 Gene.
Cell ( IF 45.5 ) Pub Date : 2018-Feb-22 , DOI: 10.1016/j.cell.2018.01.012
X Shawn Liu 1 , Hao Wu 2 , Marine Krzisch 1 , Xuebing Wu 1 , John Graef 3 , Julien Muffat 1 , Denes Hnisz 1 , Charles H Li 4 , Bingbing Yuan 1 , Chuanyun Xu 1 , Yun Li 1 , Dan Vershkov 5 , Angela Cacace 3 , Richard A Young 4 , Rudolf Jaenisch 4
Affiliation  

Fragile X syndrome (FXS), the most common genetic form of intellectual disability in males, is caused by silencing of the FMR1 gene associated with hypermethylation of the CGG expansion mutation in the 5' UTR of FMR1 in FXS patients. Here, we applied recently developed DNA methylation editing tools to reverse this hypermethylation event. Targeted demethylation of the CGG expansion by dCas9-Tet1/single guide RNA (sgRNA) switched the heterochromatin status of the upstream FMR1 promoter to an active chromatin state, restoring a persistent expression of FMR1 in FXS iPSCs. Neurons derived from methylation-edited FXS iPSCs rescued the electrophysiological abnormalities and restored a wild-type phenotype upon the mutant neurons. FMR1 expression in edited neurons was maintained in vivo after engrafting into the mouse brain. Finally, demethylation of the CGG repeats in post-mitotic FXS neurons also reactivated FMR1. Our data establish that demethylation of the CGG expansion is sufficient for FMR1 reactivation, suggesting potential therapeutic strategies for FXS.

中文翻译:


通过 FMR1 基因的 DNA 甲基化编辑拯救脆性 X 综合征神经元。



脆性 X 综合征 (FXS) 是男性智力障碍最常见的遗传形式,是由 FMR1 基因沉默引起的,该基因沉默与 FXS 患者中 FMR1 5' UTR 中 CGG 扩展突变的高甲基化相关。在这里,我们应用最近开发的 DNA 甲基化编辑工具来逆转这种超甲基化事件。 dCas9-Tet1/单引导 RNA (sgRNA) 对 CGG 扩增进行靶向去甲基化,将上游 FMR1 启动子的异染色质状态转变为活性染色质状态,从而恢复 FMR1 在 FXS iPSC 中的持续表达。源自甲基化编辑的 FXS iPSC 的神经元挽救了电生理学异常,并恢复了突变神经元的野生型表型。移植到小鼠大脑后,编辑过的神经元中的 FMR1 表达在体内得以维持。最后,有丝分裂后 FXS 神经元中 CGG 重复序列的去甲基化也重新激活了 FMR1。我们的数据表明,CGG 扩增的去甲基化足以重新激活 FMR1,这表明 FXS 的潜在治疗策略。
更新日期:2018-02-15
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