Position-dependent effects of hnRNP A1/A2 in SMN1/2 exon7 splicing.
Position-dependent effects of hnRNP A1/A2 in SMN1/2 exon7 splicing.
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DOI：
10.1016/j.bbagrm.2022.194875
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发表时间：
2022-10
期刊：
影响因子：
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通讯作者：
Jiaying Qiu;Ruobing Qu;Mengsi Lin;Jian Xu;Qingwen Zhu;Zhenyu Zhang;Junjie Sun
中科院分区：
文献类型：
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作者：
Jiaying Qiu;Ruobing Qu;Mengsi Lin;Jian Xu;Qingwen Zhu;Zhenyu Zhang;Junjie Sun
异质核核糖核蛋白A1和A2（hnRNP A1/2）是一种广泛表达的RNA结合蛋白，已知其结合内含子或外显子剪接沉默子。hnRNPA1/2与运动神经元存活基因（SMN 1/2）外显子7及其侧翼序列的结合强烈抑制了外显子7的包含，该外显子7导致脊髓性肌萎缩症，这是一种常见的遗传性疾病。然而，hnRNP A1/2在远离外显子7一侧的作用尚不清楚。我们利用反义寡核苷酸在SMN1/2基因内含子7的3 ′剪接位点（SS）附近钓取了一个内含子剪接增强子（ISE）。突变鉴定ISE的有效基序为"UAGUAGG"，结合RNA下拉和蛋白过表达，我们证明hnRNP A1/2与ISE结合促进SMN 1/2外显子7的包含。利用MS2-tethering array和"UAGGGU" motif walking进一步揭示了hnRNP A1/2对SMN1/2外显子7剪接的影响是位置依赖性的：当hnRNP A1/2与内含子7的5 ′ SS近端结合时，外显子7包含被抑制，当其与内含子7的3 ′ SS近端结合时，外显子7包含被促进。这些数据为SMN1/2的剪接调控机制提供了新的见解。
Heterogeneous nuclear ribonucleoprotein A1 and A2 (hnRNP A1/2) is a ubiquitously expressed RNA binding protein known to bind intronic or exonic splicing silencer. Binding of hnRNP A1/2 tosurvival of motor neurongene(SMN1/2) exon 7 and flanking sequences strongly inhibits the inclusion of exon 7, which causes spinal muscular atrophy, a common genetic disorder. However, the role of hnRNP A1/2 on the side away from exon 7 is unclear. Here using antisense oligonucleotides, we fished an intronic splicing enhancer (ISE) near the 3′-splice site (SS) of intron 7 ofSMN1/2. Mutagenesis identified the efficient motif of the ISE as “UAGUAGG”, coupled with RNA pull down and protein overexpression, we proved that hnRNP A1/2 binding to the ISE promotes the inclusion ofSMN1/2exon 7. Using MS2-tethering array and “UAGGGU” motif walking, we further uncovered that effects of hnRNP A1/2 onSMN1/2exon 7 splicing are position-dependent: exon 7 inclusion is inhibited when hnRNP A1/2 binds proximal to the 5′SS of intron 7, promoted when its binds proximal to the 3′SS. These data provide new insights into the splicing regulatory mechanism ofSMN1/2.