Molecular Mechanisms and Signal-Induced Regulation of Alternative Splicing

项目摘要 该项目的最终目标是了解RNA加工的分子机制， 由信号通路控制，以及这种调节对细胞功能的影响。的具体重点 目前的资助期是在T细胞活化过程中调节替代性前mRNA剪接。替代 剪接是基因调控的基本和普遍存在的机制，其允许多样化， 在不同的细胞类型或环境条件下控制蛋白质表达。特别是， 确定了信号诱导的选择性剪接在神经元活动，营养感测， 肿瘤发生和免疫功能。然而，关于信号通路如何控制替代选择的机制观点， 仍然缺乏剪接。T细胞活化为复杂的细胞应答提供了极好的模型系统， 因为多个信号传导途径在抗原接合的下游被触发并且单独地起作用， 协同地诱导T细胞效应子功能。重要的是，已知有几百个基因 选择性剪接响应T细胞活化;然而，仍有待确定这些基因是如何被激活的。 调控，哪些基因是由重叠机制/途径共同调控的，以及最终的调控机制是什么？ 功能性的结果是这样的规则。这一建议将解决这些悬而未决的问题的信号- 通过利用最近开发的方法和系统来确定：（1） 特异性信号传导途径导致T细胞抗原刺激后可变剪接的变化， 以及连接每个信号通路及其各自剪接的RNA结合蛋白（RBP） （2）RBP和/或细胞信号传导调节细胞中特定转换的分子机制。 剪接体组装指导同种型表达;（3）选择性剪接对细胞功能的影响 通过调节相关激酶的同种型表达进行信号传导;以及（4）如何调节替代激酶的表达。 剪接与其它RNA生物发生事件如转录和3 ′末端加工相协调。 总之，这些研究将提供新的见解，关于信号和剪接的相互作用，在塑造 T细胞活化过程中的细胞功能。由于在T细胞活化时诱导的信号传导途径也是 在与细胞生长、代谢和癌症相关的途径中起作用，这些研究中获得的见解将是 广泛适用于远远超出免疫系统的许多细胞反应。此外，研究 本文提出的方法将揭示细胞调节的分子机制的新范例 剪接体组装，以及RBP如何协调控制剪接沿着RNA加工中的其他步骤。 因此，这些实验的结果将大大增加对控制细胞凋亡的机制的理解。 选择性剪接是调节人类所有细胞基因表达的一个重要和普遍存在的过程， 类型

Project Summary The ultimate goal of this project is to understand the molecular mechanisms by which RNA processing is controlled by signaling pathways, and the impact of this regulation on cellular function. The specific focus of the current funding period is on regulation of alternative pre-mRNA splicing during T cell activation. Alternative splicing is an essential and ubiquitous mechanism of gene regulation, which allows for the diversification and control of protein expression in distinct cell-types or environmental conditions. In particular, it is well established that signal-induced alternative splicing is pervasive during neuronal activity, nutrient sensing, oncogenesis and immune function. However, a mechanistic view of how signaling pathways control alternative splicing remains lacking. T cell activation provides an excellent a model system for complex cellular responses, in that multiple signaling pathways are triggered downstream of antigen engagement and act, individually and cooperatively, to induce T cell effector functions. Importantly, several hundred genes are known to undergo alternative splicing in response to T cell activation; however, it remains to be determined how these genes are regulated, which genes are co-regulated by overlapping mechanisms/pathways, and what the ultimate functional consequence is of such regulation. This proposal will address these unanswered questions of signal- induced alternative splicing by leveraging recently developed methodologies and systems to determine: (1) the specific signaling pathways that lead to changes in alternative splicing following antigen stimulation of T cells, and the RNA binding proteins (RBPs) that connect each signaling pathway with their respective splicing targets; (2) the molecular mechanisms by which RBPs and/or cell signaling regulate specific transitions in spliceosome assembly to direct isoform expression; (3) the functional impact of alternative splicing on cell signaling through regulating isoform expression of related kinases; and (4) how regulation of alternative splicing is coordinated with other RNA biogenesis events such as transcription and 3' end processing. Together these studies will provide novel insight regarding the interplay of signaling and splicing in shaping cellular function during T cell activation. Since the signaling pathways induced upon T cell activation are also functional in pathways related to cell growth, metabolism and cancer, the insight gained in these studies will be broadly applicable to numerous cellular responses far beyond the immune system. In addition, the studies proposed here will reveal new paradigms regarding the molecular mechanisms by which cells regulate spliceosome assembly, and how RBPs coordinately control splicing along with other steps in RNA processing. Results from these experiments will thus significantly increase understanding of the mechanisms that control alternative splicing, an essential and ubiquitous process for regulating gene expression across all human cell types.