Double synonymous mutations in exon 9 of the Cullin3 gene restore exon inclusion by abolishing hnRNPs inhibition.
Double synonymous mutations in exon 9 of the Cullin3 gene restore exon inclusion by abolishing hnRNPs inhibition.
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Cullin3 基因外显子 9 中的双同义突变通过消除 hnRNP 抑制来恢复外显子包含。
DOI：
10.1093/hmg/ddac148
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发表时间：
2022-07
期刊：
影响因子：
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通讯作者：
赵向忠
中科院分区：
文献类型：
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作者：
刘志英;隋爱华;望赛;崔莉;辛卿;张瑞晓;韩玥;邵乐平;赵向忠
与假性醛固酮减少症II型（PHA II）相关的Cullin 3基因外显子9的所有突变都不同程度地导致外显子跳跃，但这种异常剪接的具体分子机制仍不清楚。本研究的目的是研究两个同义剪接事件c.1221A > G（p.Glu407Glu）和c.1236G > A（p.Leu412Leu）的调控机制，并发现通过靶向潜在的调控位点来纠正这种异常剪接的治疗策略。通过一系列RNA pulldown、银染、western blotting、small interfering RNA knockdown、体外过表达和单或双位点定向诱变实验，我们首先探索了由CUL3基因突变引起的外显子9跳跃的发病机制，并证实了与同义的c.1221A > G和c.1236G > A突变是核内不均一的核糖核蛋白。此外，我们证实了引入另一个同义突变c.1224A > G（A18G），显著挽救了由c.1221A > G和c.1236G > A引起的异常剪接，突出了治疗PHA II的治疗潜力。
All mutations in exon 9 of the Cullin3 gene associated with pseudohypoaldosteronism type II (PHA II) contribute to exon skipping to different degrees, but the specific molecular mechanism of this aberrant splicing is still unclear. The aims of this study were to investigate the regulatory mechanism underlying two synonymous splicing events, c.1221A > G (p. Glu407Glu) and c.1236G > A (p. Leu412Leu), and to discover a therapeutic strategy for correcting this aberrant splicing by targeting potential regulatory sites. Through a series of RNA pulldown, silver staining, western blotting, small interfering RNA knockdown, in vitro overexpression and single or double site-directed mutagenesis experiments, we first explored the pathogenesis of exon 9 skipping caused by mutations in the CUL3 gene and verified that the main splicing regulators associated with the synonymous c.1221A > G and c.1236G > A mutations were heterogeneous nuclear ribonucleoproteins. In addition, we verified that introducing another synonymous mutation, c.1224A > G (A18G), significantly rescued the abnormal splicing caused by c.1221A > G and c.1236G > A, highlighting the therapeutic potential for the treatment of PHA II.