Department of Drug Discovery Medicine, Kyoto University Graduate School of Medicine, Kyoto, Japan.
Department of Anatomy and Developmental Biology, Kyoto University Graduate School of Medicine, Kyoto, Japan.
Nat Commun. 2021 Jul 23;12(1):4507. doi: 10.1038/s41467-021-24705-5.
Approximately half of genetic disease-associated mutations cause aberrant splicing. However, a widely applicable therapeutic strategy to splicing diseases is yet to be developed. Here, we analyze the mechanism whereby IKBKAP-familial dysautonomia (FD) exon 20 inclusion is specifically promoted by a small molecule splice modulator, RECTAS, even though IKBKAP-FD exon 20 has a suboptimal 5' splice site due to the IVS20 + 6 T > C mutation. Knockdown experiments reveal that exon 20 inclusion is suppressed in the absence of serine/arginine-rich splicing factor 6 (SRSF6) binding to an intronic splicing enhancer in intron 20. We show that RECTAS directly interacts with CDC-like kinases (CLKs) and enhances SRSF6 phosphorylation. Consistently, exon 20 splicing is bidirectionally manipulated by targeting cellular CLK activity with RECTAS versus CLK inhibitors. The therapeutic potential of RECTAS is validated in multiple FD disease models. Our study indicates that small synthetic molecules affecting phosphorylation state of SRSFs is available as a new therapeutic modality for mechanism-oriented precision medicine of splicing diseases.
大约一半的遗传疾病相关突变导致异常剪接。然而，一种广泛适用的治疗剪接疾病的策略尚未开发出来。在这里，我们分析了小分子剪接调节剂 RECTAS 特异性促进 IKBKAP-家族性自主神经异常（FD）外显子 20 包含的机制，尽管由于 IVS20+6T>C 突变，IKBKAP-FD 外显子 20 具有次优的 5'剪接位点。敲低实验表明，在缺乏丝氨酸/精氨酸丰富的剪接因子 6（SRSF6）与内含子 20 中的内含子剪接增强子结合的情况下，外显子 20 包含被抑制。我们表明，RECTAS 直接与细胞周期蛋白样激酶（CLKs）相互作用，并增强 SRSF6 的磷酸化。一致地，通过用 RECTAS 与 CLK 抑制剂靶向细胞 CLK 活性，外显子 20 剪接被双向操纵。RECTAS 在多种 FD 疾病模型中的治疗潜力得到了验证。我们的研究表明，影响 SRSFs 磷酸化状态的小分子合成物可作为剪接疾病的机制定向精准医学的一种新治疗模式。
