Dissecting the Mechanisms of Quaking-Regulated RNA Processing

摘要： 这项研究计划的总体目标是确定QKI家族的Quaking-5亚型（QKI 5）如何与人类的免疫缺陷病毒（HIV）结合。 RNA结合蛋白调节基因表达。为了实现这一目标，我们将确定如何监管QKI 5， 它如何执行调节功能，以及这些活动对整个基因的影响 表达式程序追求这一目标是我实验室的主要使命，也将是一个引人入胜的课题 在此基础上，我将发展我的实验室，培养博士后和研究生。QKI是一种保守的调节因子， 发育和组织特异性基因表达，但其功能的分子机制， 执行的研究仍然不足，特别是对于QKI 5同种型。它的重要性是由多个 在心脏病、癌症、精神分裂症和其他严重疾病中涉及QKI功能失调的观察结果 条件，以及我们最近发现它调节谱系特异性基因表达。因此，该RBP可以 代表了合理的，基于RNA的药物设计的治疗漏洞，并告诉我们如何最早的 细胞命运的决定。尽管有这种可能性，但在理解方面存在重大差距，必须 在这成为现实之前，这就要求我们通过 哪一个单独的靶RNA和整个转录组被调节。因此，存在着重大的知识差距 在理解QKI 5用于执行其前mRNA剪接的基本机制时， RNA加工中的定位和稳定性/衰变功能。这项研究计划将利用我们的分子 遗传学，生物化学和基因组学专业知识，以解决有关这一主题的知识的关键差距：1）如何 QKI 5的上游调控影响其调节前mRNA剪接的能力，以及2）QKI 5如何直接和 间接调节RNA剪接。解决这些关键差距将解决上游、中间和 QKI法规对下游的影响。我们将使用人类和小鼠的细胞，以及细胞提取物， 进行这些研究。我们认为QKI 5的结合（以及剪接活性）受到以下因素的影响：a）竞争 前体mRNA底物和lncRNA之间的相互作用，和B）蛋白质-蛋白质相互作用。此外，我们建议， 由QKI 5调控的剪接与转录本定位和稳定性相关，QKI 5影响核心转录因子的表达。 剪接机器这些研究将产生基础数据，为更广泛的科学研究提供信息。 RBP如何被调控和调控基因表达。总的来说，这些发现将显示 复杂的多步骤RNA加工如何整合到分子信息的整体流程中， 维持细胞内环境稳定。

Abstract: The overall goal of this research program is to determine how the Quaking-5 isoform (QKI5) of the QKI family of RNA binding proteins regulates gene expression. To achieve this goal, we will determine how QKI5 is regulated, how it executes regulatory functions, and what the consequences of these activities are on the overall gene expression program. Pursuing this goal is the primary mission of my laboratory, and will be an engaging topic upon which I will develop my lab and train post-docs and graduate students. QKI is a conserved regulator of development and tissue-specific gene expression, yet the molecular mechanisms through which its functions are executed remain understudied, particularly for the QKI5 isoform. Its importance is underscored by multiple observations implicating dysregulated QKI function in heart disease, cancer, schizophrenia, and other serious conditions, and our recent discovery that it regulates lineage-specific gene expression. Therefore, this RBP may represent a therapeutic vulnerability for rational, RNA-based drug design, and inform us as to how the earliest cell fate decisions are made. Despite this possibility, there are significant gaps in understanding that must be filled prior to this becoming a reality. This requires us to illuminate the precise molecular mechanisms through which individual target RNAs and the entire transcriptome are regulated. Hence, a major knowledge gap exists in understanding the fundamental mechanisms used by QKI5 to execute its pre-mRNA splicing, transcript localization, and stability/decay functions in RNA processing. This research program will leverage our molecular genetics, biochemical, and genomics expertise to address key gaps in knowledge pertaining to this topic: 1) how upstream regulation of QKI5 impacts its ability to regulate pre-mRNA splicing, and 2) how QKI5 directly and indirectly regulates RNA splicing. Addressing these key gaps will tackle the upstream, intermediate, and downstream impacts on, of, and by QKI regulation. We will use human and mouse cells, and cell extracts to conduct these studies. We propose that QKI5 binding (and thus splicing activity) is influenced by a) competition between pre-mRNA substrates and lncRNAs, and b) protein-protein interactions. Moreover, we propose that regulated splicing by QKI5 is coupled to transcript localization and stability, and that QKI5 influences the core spliceosome machinery. These studies will yield fundamental data that will inform the broader scientific community as to how RBPs can be regulated and regulate gene expression. Overall, these findings will show how complex, multi-step RNA processing is integrated into the overall flow of molecular information in order to maintain cellular homeostasis.