The Jackson Laboratory for Genomic Medicine, Farmington, CT, USA; Graduate Program in Genetics and Development, UConn Health, Farmington, CT, USA.
The Jackson Laboratory for Genomic Medicine, Farmington, CT, USA.
Mol Cell. 2020 Nov 19;80(4):648-665.e9. doi: 10.1016/j.molcel.2020.10.019. Epub 2020 Nov 10.
The RNA isoform repertoire is regulated by splicing factor (SF) expression, and alterations in SF levels are associated with disease. SFs contain ultraconserved poison exon (PE) sequences that exhibit greater identity across species than nearby coding exons, but their physiological role and molecular regulation is incompletely understood. We show that PEs in serine-arginine-rich (SR) proteins, a family of 14 essential SFs, are differentially spliced during induced pluripotent stem cell (iPSC) differentiation and in tumors versus normal tissues. We uncover an extensive cross-regulatory network of SR proteins controlling their expression via alternative splicing coupled to nonsense-mediated decay. We define sequences that regulate PE inclusion and protein expression of the oncogenic SF TRA2β using an RNA-targeting CRISPR screen. We demonstrate location dependency of RS domain activity on regulation of TRA2β-PE using CRISPR artificial SFs. Finally, we develop splice-switching antisense oligonucleotides to reverse the increased skipping of TRA2β-PE detected in breast tumors, altering breast cancer cell viability, proliferation, and migration.
RNA 异构体谱受剪接因子 (SF) 表达调控，SF 水平的改变与疾病有关。SF 包含超保守的毒蕈碱外显子 (PE) 序列，这些序列在物种间的同一性高于附近的编码外显子，但它们的生理功能和分子调控机制尚不完全清楚。我们表明，在诱导多能干细胞 (iPSC) 分化过程中以及在肿瘤与正常组织中，丝氨酸-精氨酸丰富 (SR) 蛋白家族中的 14 种必需 SF 中的 PE 是有差异剪接的。我们发现了一个广泛的 SR 蛋白交叉调控网络，通过与无意义介导的衰变偶联的选择性剪接来控制它们的表达。我们使用靶向 RNA 的 CRISPR 筛选来定义调节致癌性 SF TRA2β 的 PE 包含和蛋白质表达的序列。我们证明了 RS 结构域活性对 TRA2β-PE 调节的位置依赖性，使用 CRISPR 人工 SF。最后，我们开发了剪接转换反义寡核苷酸来逆转在乳腺癌中检测到的 TRA2β-PE 过度跳跃，改变乳腺癌细胞活力、增殖和迁移。
