Regulation of alternative splicing during epithelial-mesenchymal transition

上皮间质转化过程中选择性剪接的调节

• 批准号: 10364651
• 负责人: Chonghui Cheng
• 金额: $ 40万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2024-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10364651
• 关键词: Address; Alternative Splicing; Bioinformatics; Biological; CD44 gene; Cells; Cellular biology; Development; Developmental Process; Disease; Epithelial; Fibrosis; Genes; Goals; Mammals; Mesenchymal; Messenger RNA; Molecular; Neoplasm Metastasis; Phenotype; Phosphotransferases; Protein Isoforms; RNA; RNA Splicing; RNA-Binding Proteins; Regulation; Research; Signal Transduction; System; Time; Tissues; Transcript; Work; causal variant; cell motility; experimental analysis; insight; mRNA Precursor; programs

ABSTRACT Alternative splicing of pre-mRNA enables a gene to give rise to multiple distinct mRNA transcripts, yielding protein isoforms with different, even opposing, functions. The long-term goal of our research is to understand the molecular mechanisms and function of alternative splicing on biological activities. The focus of this proposal is to investigate the regulation of alternative splicing in epithelial-mesenchymal transition (EMT). EMT is an essential developmental process that allows cells to change from a tightly packed cobble-stone-like epithelial cellular state to a motile and spindle-shaped mesenchymal cellular state. When abnormally activated, EMT promotes many types of diseases, including tissue fibrosis and cancer metastasis. Through working at the intersection of RNA splicing and cell biology, my lab has made several important findings connecting RNA regulation and EMT. Our work revealed, for the first time, that splice isoform switching of the CD44 gene causally controls EMT. By manipulating CD44 alternative splicing, we were able to convert cells between the epithelial and mesenchymal states. We have also identified the RNA-binding protein (RBP) hnRNPM as a critical splicing factor that promotes EMT through the regulation of alternative splicing. These results suggest that RNA splicing regulation could serve as an important mechanism that provides cellular plasticity. By shifting the programs of alternative splicing, cells are able to convert between the epithelial and mesenchymal cellular states. This capacity of reversing phenotypes is important for normal developmental EMT, as well as for cancer metastasis. Major new discoveries are necessary to fully understand this phenomenon and its underlying mechanisms. Our proposed research program is focused on (1) determining whether splice isoform switching acts as a prevalent mechanism that drives EMT or, if it is largely a phenomenon of byproducts; (2) understanding how RBPs precisely control alternative splicing during EMT; and (3) dissecting how signaling cascades elicit signals to RBPs and trigger alternative splicing changes during EMT. We have made major efforts in the past several years to build up experimental systems and gain expertise to help carry out our proposed studies. These efforts include large-scale RNA profiling of alternative splicing in multiple EMT systems, bioinformatics and experimental analysis of RBPs, as well as our recently completed kinase screen for EMT-associated alternative splicing alterations. Accomplishing the proposed work will provide new insights into our understanding of the regulatory mechanisms of alternative splicing, thus contributing to biological relevance in normal development and diseases.

抽象的 前 mRNA 的选择性剪接使基因能够产生多个不同的 mRNA 转录本， 产生具有不同甚至相反功能的蛋白质亚型。我们研究的长期目标是 了解选择性剪接对生物活性的分子机制和功能。重点是 该提案旨在研究选择性剪接在上皮间质转化（EMT）中的调控。 EMT 是一个重要的发育过程，使细胞从紧密堆积的鹅卵石状转变为 上皮细胞状态转变为活动的纺锤形间充质细胞状态。当异常启动时， EMT 会促进多种疾病，包括组织纤维化和癌症转移。通过在 RNA 剪接和细胞生物学的交叉点，我的实验室在连接 RNA 方面取得了多项重要发现 监管和 EMT。我们的工作首次揭示了 CD44 基因的剪接异构体转换与 控制 EMT。通过操纵 CD44 选择性剪接，我们能够在上皮细胞和上皮细胞之间转换细胞。 和间充质状态。我们还确定了 RNA 结合蛋白 (RBP) hnRNPM 作为关键剪接 通过调节选择性剪接促进 EMT 的因素。这些结果表明 RNA 剪接 调节可以作为提供细胞可塑性的重要机制。通过改变程序 通过选择性剪接，细胞能够在上皮细胞和间充质细胞状态之间转换。这 逆转表型的能力对于正常发育 EMT 以及癌症转移非常重要。 为了充分理解这种现象及其潜在机制，需要重大的新发现。我们的 拟议的研究计划侧重于（1）确定剪接异构体转换是否作为一种普遍的 驱动 EMT 的机制，或者，如果它主要是一种副产品现象； (2)了解RBP如何 EMT期间精确控制选择性剪接； (3) 剖析信号级联如何向 RBP 引出信号 并在 EMT 期间触发选择性剪接变化。在过去的几年里，我们做出了巨大的努力 建立实验系统并获得专业知识以帮助开展我们提出的研究。这些努力包括 多个 EMT 系统、生物信息学和实验中选择性剪接的大规模 RNA 分析 RBP 分析，以及我们最近完成的 EMT 相关选择性剪接激酶筛选 变更。完成拟议的工作将为我们对监管的理解提供新的见解 选择性剪接机制，从而有助于正常发育和疾病的生物学相关性。