Post-Transcriptional controls in Mammalian Erythroid Differentiation

描述（由申请人提供）：该基金的长期目标是建立对哺乳动物红细胞分化至关重要的转录后控制的全面理解。红细胞分化的标志是一组戏剧性的形态和功能的变化，这个过程中发生的转录沉默的环境。由于这个原因，红细胞分化严重依赖于转录后控制。RNA结合蛋白（RBP）是转录后调控的主要调节因子，影响细胞核中的转录物加工以及细胞质中mRNA的稳定性和功能。RBP在其结构、结合特异性和细胞区室化方面变化很大，并且可以通过直接作用和/或通过效应复合物的募集来调节RNA功能。RBP，其mRNA的目标，和相应的转录后控制的鉴定，推进了我们对许多发育过程的理解，并揭示了意想不到的病理生理途径。通过定义转录后控制红细胞分化，我们将扩大我们的理解，遗传性和获得性红细胞生成障碍，并建立一个模板，在其他系统中进行类似的调查。为了实现这一目标，我们将进行一种新的和最先进的全转录组分析，以揭示伴随红细胞终末分化动态过程的全方位mRNA-蛋白质（mRNP）相互作用。这种方法将生物化学方法与专门为复杂RNA-蛋白质相互作用的全面描述而设计的创新信息管道相结合。这种无偏见的分析将结合一系列有针对性的深入机制研究，重点是两组mRNA/蛋白质相互作用，我们已经确定发挥核心作用，在红细胞分化。这些互补方法的组合输出将在这个发育稳健且临床相关的模型系统中建立独特的转录后控制网络。这项建议包括三个具体目标。目标1.描述聚C结合蛋白β 1和β 2作为红细胞分化的转录后整合子的作用。目标2.确定polyA结合蛋白PABPC在定义和维持红系转录组中的关键作用。目标3。绘制红系细胞中mRNA/蛋白质相互作用的全局结构，并定义这些相互作用在分化过程中的动态性质。将转录组范围的分析与有针对性的机制研究相结合，将有力地获得构成红细胞分化关键决定因素的RNA/蛋白质相互作用的复杂阵列。这些研究的成功将从根本上改变我们对这一深入研究的途径的理解;作为研究其他系统中转录后控制的原型，并为未来的诊断和治疗创新提供新的靶点。

DESCRIPTION (provided by applicant): The long-term goal of this grant is to establish a comprehensive understanding of post-transcriptional controls critical to mammalian erythroid differentiation. Erythroid differentiation is marked by a set of dramatic morphologic and functional changes, much of this process occurring in a transcriptionally-silent environment. For this reason erythroid differentiation is heavily dependent on post-transcriptional controls. RNA binding proteins (RBPs), the major regulators of post-transcriptional controls, impact on transcript processing in the nucleus as well as the stability and function of the mRNA in the cytoplasm. RBPs vary widely in their structures, binding specificities, and cellular compartmentalization and can modulate RNA functions by direct actions and/or via the recruitment of effector complexes. Identification of RBPs, their mRNA targets, and corresponding post-transcriptional controls have advanced our understanding of numerous developmental processes and have revealed unanticipated pathophysiologic pathways. By defining post-transcriptional controls in erythroid differentiation, we will expand our understanding of inherited and acquired disorders of erythropoiesis and establish a template for similar investigations in other systems. To achieve this goal, we will carry out a novel and state-of-the-art transcriptome-wide analyses to reveal the full range of mRNA-protein (mRNP) interactions that accompany the dynamic process of erythroid terminal differentiation. This approach combines biochemical methodologies with innovative informatic pipelines specifically designed for comprehensive descriptions of complex RNA-protein interactions. This unbiased analysis will be combined with a series of targeted in-depth mechanistic studies that focus on two sets of mRNA/protein interactions that we have identified to play central roles in erythroid differentiation. The combined output of these complementing approaches will establish the unique network of post-transcriptional controls in this developmentally robust and clinically relevant model system. This proposal encompasses three Specific Aims. Aim 1. Characterize the roles of the polyC-binding proteins, �1 and �2, as post- transcriptional integrators of erythroid differentiation. Aim 2. Identify the critical role(s) of the polyA binding protein, PABPC, in defining and sustaining the erythroid transcriptome. Aim 3. Map the global structure of mRNA/protein interactions in erythroid cells and define the dynamic nature of these interactions in the differentiation process. Combining transcriptome-wide analyses with targeted mechanistic studies will generate a powerful access to the complex array of RNA/protein interactions that constitute critical determinants of erythroid differentiation. Success in these studies will fundamentally alter our understanding of this intensively studied pathway; serve as a prototype for investigations of post- transcriptional controls in other systems, and present novel targets fo future diagnostic and therapeutic innovations.