Koch Institute for Integrative Cancer Research, Massachusetts Institute of Technology, Cambridge, Massachusetts, USA.
PLoS Genet. 2011 Aug;7(8):e1002218. doi: 10.1371/journal.pgen.1002218. Epub 2011 Aug 18.
Epithelial-mesenchymal transition (EMT), a mechanism important for embryonic development, plays a critical role during malignant transformation. While much is known about transcriptional regulation of EMT, alternative splicing of several genes has also been correlated with EMT progression, but the extent of splicing changes and their contributions to the morphological conversion accompanying EMT have not been investigated comprehensively. Using an established cell culture model and RNA-Seq analyses, we determined an alternative splicing signature for EMT. Genes encoding key drivers of EMT-dependent changes in cell phenotype, such as actin cytoskeleton remodeling, regulation of cell-cell junction formation, and regulation of cell migration, were enriched among EMT-associated alternatively splicing events. Our analysis suggested that most EMT-associated alternative splicing events are regulated by one or more members of the RBFOX, MBNL, CELF, hnRNP, or ESRP classes of splicing factors. The EMT alternative splicing signature was confirmed in human breast cancer cell lines, which could be classified into basal and luminal subtypes based exclusively on their EMT-associated splicing pattern. Expression of EMT-associated alternative mRNA transcripts was also observed in primary breast cancer samples, indicating that EMT-dependent splicing changes occur commonly in human tumors. The functional significance of EMT-associated alternative splicing was tested by expression of the epithelial-specific splicing factor ESRP1 or by depletion of RBFOX2 in mesenchymal cells, both of which elicited significant changes in cell morphology and motility towards an epithelial phenotype, suggesting that splicing regulation alone can drive critical aspects of EMT-associated phenotypic changes. The molecular description obtained here may aid in the development of new diagnostic and prognostic markers for analysis of breast cancer progression.
上皮-间质转化（EMT）是胚胎发育过程中重要的机制，在恶性转化过程中起着关键作用。虽然已经了解了 EMT 的转录调控，但几个基因的选择性剪接也与 EMT 进展相关，但剪接变化的程度及其对伴随 EMT 的形态转化的贡献尚未得到全面研究。我们使用已建立的细胞培养模型和 RNA-Seq 分析，确定了 EMT 的选择性剪接特征。编码 EMT 依赖性细胞表型变化的关键驱动基因，如肌动蛋白细胞骨架重塑、细胞-细胞连接形成的调节以及细胞迁移的调节，在 EMT 相关的选择性剪接事件中富集。我们的分析表明，大多数 EMT 相关的选择性剪接事件是由 RBFOX、MBNL、CELF、hnRNP 或 ESRP 类剪接因子的一个或多个成员调节的。EMT 选择性剪接特征在人乳腺癌细胞系中得到了验证，这些细胞系可以仅根据其 EMT 相关的剪接模式分为基底和腔型。在原发性乳腺癌样本中也观察到 EMT 相关的选择性 mRNA 转录本的表达，表明 EMT 依赖性剪接变化在人类肿瘤中普遍发生。通过表达上皮特异性剪接因子 ESRP1 或在间质细胞中耗尽 RBFOX2 来测试 EMT 相关的选择性剪接的功能意义，这两种方法都引起了细胞形态和向上皮表型的运动的显著变化，表明剪接调节本身可以驱动 EMT 相关表型变化的关键方面。这里获得的分子描述可能有助于开发新的诊断和预后标志物，用于分析乳腺癌的进展。
