Department of Biology and Biotechnology, 'Charles Darwin', Sapienza University of Rome, Rome, Italy.
CNR Institute of Molecular Biology and Pathology (IBPM), Rome, Italy.
EMBO Mol Med. 2020 Aug 7;12(8):e12063. doi: 10.15252/emmm.202012063. Epub 2020 Jun 29.
Exon skipping is an effective strategy for the treatment of many Duchenne Muscular Dystrophy (DMD) mutations. Natural exon skipping observed in several DMD cases can help in identifying novel therapeutic tools. Here, we show a DMD study case where the lack of a splicing factor (Celf2a), which results in exon skipping and dystrophin rescue, is due to a maternally inherited trans-generational epigenetic silencing. We found that the study case and his mother express a repressive long non-coding RNA, DUXAP8, whose presence correlates with silencing of the Celf2a coding region. We also demonstrate that DUXAP8 expression is lost upon cell reprogramming and that, upon induction of iPSCs into myoblasts, Celf2a expression is recovered leading to the loss of exon skipping and loss of dystrophin synthesis. Finally, CRISPR/Cas9 inactivation of the splicing factor Celf2a was proven to ameliorate the pathological state in other DMD backgrounds establishing Celf2a ablation or inactivation as a novel therapeutic approach for the treatment of Duchenne Muscular Dystrophy.
外显子跳跃是治疗许多杜氏肌营养不良症（DMD）突变的有效策略。在几种 DMD 病例中观察到的自然外显子跳跃有助于鉴定新的治疗工具。在这里，我们展示了一个 DMD 病例研究，其中一个 splicing 因子（Celf2a）的缺失导致外显子跳跃和肌营养不良蛋白的拯救，这是由于母系遗传的跨代表观遗传沉默。我们发现，该病例及其母亲表达一种抑制性的长非编码 RNA，DUXAP8，其存在与 Celf2a 编码区域的沉默相关。我们还证明，细胞重编程后 DUXAP8 的表达会丢失，并且在诱导 iPSCs 分化为成肌细胞后，Celf2a 的表达会恢复，导致外显子跳跃丢失和肌营养不良蛋白合成丢失。最后，CRISPR/Cas9 敲除剪接因子 Celf2a 被证明可以改善其他 DMD 背景下的病理状态，确立了 Celf2a 缺失或失活作为治疗杜氏肌营养不良症的新治疗方法。
