The landscape of alternative polyadenylation during EMT and its regulation by the RNA-binding protein Quaking

Epithelial-mesenchymal transition (EMT) plays important roles in tumour progression and is orchestrated by dynamic changes in gene expression. While it is well established that post-transcriptional regulation plays a significant role in EMT, the extent of alternative polyadenylation (APA) during EMT has not yet been explored. Using 3’ end anchored RNA sequencing, we mapped the alternative polyadenylation (APA) landscape following Transforming Growth Factor (TGF)-β-mediated induction of EMT in human mammary epithelial cells and found APA generally causes 3’UTR lengthening during this cell state transition. Investigation of potential mediators of APA indicated the RNA-binding protein Quaking (QKI), a splicing factor induced during EMT, regulates a subset of events including the length of its own transcript. Analysis of QKI crosslinked immunoprecipitation (CLIP)-sequencing data identified the binding of QKI within 3’ untranslated regions (UTRs) was enriched near cleavage and polyadenylation sites. Following QKI knockdown, APA of many transcripts is altered to produce predominantly shorter 3’UTRs associated with reduced gene expression. These findings reveal the changes in APA that occur during EMT and identify a potential role for QKI in this process.

上皮间质转化（Epithelial-mesenchymal transition, EMT）在肿瘤进展中发挥关键作用，其进程受基因表达的动态变化精准调控。尽管已有研究证实转录后调控在EMT中具有重要功能，但EMT过程中可变多聚腺苷酸化（alternative polyadenylation, APA）的调控范围尚未被系统阐明。本研究采用3'端锚定RNA测序技术，对人乳腺上皮细胞中转化生长因子β（Transforming Growth Factor-β, TGF-β）诱导的EMT过程开展APA景观图谱绘制，发现在该细胞状态转变过程中，APA普遍会导致3'非翻译区（3'UTR）延长。对APA潜在调控因子的分析显示，RNA结合蛋白Quaking（QKI）——一种在EMT过程中被诱导表达的剪接因子——可调控一类APA事件，包括其自身转录本的长度变化。对QKI交联免疫沉淀测序（crosslinked immunoprecipitation-sequencing, CLIP-seq）数据的分析发现，QKI在3'UTR内的结合位点显著富集于切割与多聚腺苷酸化位点附近。在敲低QKI后，众多转录本的APA模式发生改变，主要生成更短的3'UTR，这一变化与对应基因的表达水平降低显著相关。本研究揭示了EMT过程中APA的动态变化特征，并明确了QKI在该过程中的潜在调控作用。