Splicing activation by Rbfox requires self-aggregation through its tyrosine-rich domain

To determine how the higher order assembly of Rbfox proteins affect Rbfox-dependent splicing regulation, we expressed Rbfox wildtype and its mutant protein in Flp-In™ T-REx™ 293 Rbfox2-/- cells and extracted RNA from these cells to perform RASL-seq which profiles thousands of alternative splicing event. Overall design: Flp-In™ T-REx™ 293 Cell Line (in triplicate) were transfected by Rbfox1 wildtype and its mutants, followed by Dox induction. Total RNAs were extracted and been analyzed by RASL-seq.

为探究Rbfox蛋白的高阶组装如何影响依赖于Rbfox的可变剪接（alternative splicing）调控，我们在Flp-In™ T-REx™ 293 Rbfox2-/-细胞中分别表达野生型Rbfox蛋白及其突变体，随后从这些细胞中提取RNA以开展RASL-seq实验——该技术可对数千个可变剪接事件进行图谱分析。
实验总体设计：将Rbfox1野生型及其突变体转染至Flp-In™ T-REx™ 293细胞系（设置三组生物学重复），经强力霉素（Dox）诱导表达后，提取总RNA并通过RASL-seq完成测序分析。