Molecular mechanisms underlying the establishment of cell polarity.

真核生物的一个基本特征是细胞极性的建立和维持。分子 马达通过将mRNA、蛋白质、囊泡和细胞器运输到特定位点来帮助建立极性 在细胞内。从单细胞酵母到人类的各种生物都使用mRNA定位偶联 通过翻译调节来不对称地分选蛋白质。这种现象的普遍性是 在发育中的胚胎，神经元和上皮细胞中得到了最好的说明，其中数千种mRNA在空间上被 本地化。当这个过程受到损害时，它可能导致发育和神经系统疾病。 尽管这一主题的重要性和这种现象的普遍性，我们缺乏一个机制， 了解mRNA的定位。我们理解中的一个关键差距涉及到mRNA是如何注定要 定位的mRNA被细胞识别并与非定位的mRNA区分开。除了序列 这些蛋白质存在于定位mRNA中，结合这些mRNA的蛋白质是其细胞命运的关键。这 一种蛋白质复合体负责连接定位mRNA与分子马达，并调节它们的 翻译.然而，鉴定这些关键蛋白质已被证明是极具挑战性的。就其 自然界中，mRNA定位的过程是高度动态的。因此，蛋白质-蛋白质和蛋白质-RNA 组装定位信使核糖核蛋白（mRNP）颗粒所需的相互作用很弱， 短暂的这使得使用经典的生物化学方法鉴定它们几乎是不可能的; 络合物福尔斯在纯化步骤中分解。在这个应用中，我们提出了新的策略来解决这个问题 关键的知识差距。我们建议用于这些研究的模型是具有良好特征的果蝇 黑腹产卵室在本申请的目的1中，我们提出使用邻位生物素连接来 定义传输粒子的核心组成部分，并检查这些因素之间的守恒 苍蝇和哺乳动物。在目标2中，我们提出了一个假设，即颗粒结构被称为P 体协调mRNA的定位和它们的翻译调节。

A fundamental feature of eukaryotic life is the establishment and maintenance of cellular polarity. Molecular motors help to establish polarity by transporting mRNAs, proteins, vesicles, and organelles to specific sites within the cell. A variety of organisms from the single-celled yeast to humans use mRNA localization coupled with translational regulation as a way to asymmetrically sort proteins. The prevalence of this phenomenon is best illustrated in developing embryos, neurons and epithelial, in which thousands of mRNAs are spatially localized. When this process is compromised, it can result in developmental and neurological disorders. Despite the importance of this topic and the prevalence of this phenomenon, we lack a mechanistic understanding of mRNA localization. A critical gap in our understanding pertains to how mRNAs destined for localization are recognized by the cell and distinguished from non-localizing mRNAs. In addition to sequences present within localizing mRNAs, the proteins that bind these mRNAs are key to their cellular fate. This complex of proteins is responsible for linking localizing mRNAs with molecular motors and for regulating their translation. However, identifying these critical proteins has proven to be extremely challenging. By its very nature, the process of mRNA localization is highly dynamic. Consequently, the protein-protein and protein-RNA interactions required for assembling localizing messenger ribonucleoprotein (mRNP) particles are weak and transient. This has made their identification using classical biochemical approaches almost impossible; the complex falls apart during the purification step. In this application, we propose novel strategies to address this critical knowledge gap. The model we propose to use for these studies is the well-characterized Drosophila melanogaster egg chamber. In Objective 1 of this application, we propose to use proximity biotin ligation to define the core components of transport particles and to examine the conservation of these factors between flies and mammals. In Objective 2, we propose to test the hypothesis that granular structures referred to as P bodies coordinate the localization of mRNAs with their translational regulation.