Mechanism of RNA Localization in Drosophila Development

果蝇发育中RNA定位的机制

• 批准号: 7169809
• 负责人: ELIZABETH R GAVIS
• 金额: $ 29.13万
• 依托单位: PRINCETON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2004
• 资助国家: 美国
• 起止时间: 2004-02-01 至 2008-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7169809
• 关键词: Mechanism; RNA; Localization; Drosophila; Development

DESCRIPTION (provided by applicant): Intracellular mRNA localization is an important mechanism for the spatial regulation of gene expression necessary for animal development. By restricting the distributions of developmental regulatory proteins to particular regions of oocytes and embryos, mRNA localization plays a critical role in patterning of body axes and specification of cell fates during embryonic development in a variety of organisms. In addition, RNA localization generates protein asymmetries necessary for the subsequent differentiation and function of many specialized cell types. Cis-acting signals that mediate localization have now been identified in a number of localized mRNAs. Little is known, however, about the mechanisms by which these signals are recognized specifically by cellular localization machinery and how they target their RNAs to unique intracellular locations. Restriction of Nanos protein to the posterior of the Drosophila embryo is essential for proper patterning of the anterior-posterior body axis. Nanos synthesis is limited to the posterior pole of the embryo by a combination of RNA localization and translational control. Localization of nanos mRNA to the posterior pole of the embryo generates the critical concentration Nanos protein in the posterior for abdominal development and is essential to activate nanos translation. Localization of nanos is mediated by a complex a cis-acting localization signal within its 3' untranslated region (3'UTR). Results from previous and preliminary studies indicate that cytoplasmic localization factors recognize different sequence or structural motifs within this localization signal. Using nanos as a model, the proposed work will provide insight into how complex RNA localization signals are recognized by the cellular localization machinery and how these RNA-protein interactions mediate transport and anchoring by localization pathways. More generally, these studies will shed light on mechanisms by which RNA-protein interactions provide the highly selective control of basic cellular processes needed for development, growth, and differentiation. Specific Aim 1 encompasses mutational analysis of the nanos localization signal to determine sequence and structural requirements for localization signal recognition and function. This work will be facilitated by phylogenetic analysis ofnanos 3'UTRs from ten different drosophilid species. Preliminary work has led to purification of one candidate nanos localization factor and biochemical identification of a second. Aim 2 focuses on biochemical and genetic characterization of these factors to determine their function in nanos localization. In addition, a new strategy for biochemical isolation of localization complexes is proposed. In Aim 3, a genetic screen for nanos localization factors will complement the biochemical approaches of Aim 2. Aim 4 takes advantage of a new system for GFP labeling of nanos RNA in vivo to investigate the dynamic pathway of nanos localization during oogenesis.

描述（由申请人提供）：细胞内mRNA定位是对动物发育所必需的基因表达空间调节的重要机制。通过将发育调节蛋白的分布限制为卵母细胞和胚胎的特定区域，mRNA定位在各种生物体中胚胎发育过程中的胚胎发育过程中起着至关重要的作用。此外，RNA定位会产生蛋白质不对称，这是许多专用细胞类型的随后分化和功能所必需的。现在已经在许多局部mRNA中识别出介导定位的顺式作用信号。然而，知之甚少，关于这些信号通过细胞定位机制特别识别的机制，以及它们如何将其RNA靶向独特的细胞内位置。 将纳米蛋白限制在果蝇胚胎后的后部，对于适当对前后身体轴的图案至关重要。通过RNA定位和翻译控制的组合，纳米合成仅限于胚胎的后极。纳米mRNA在胚胎的后极点定位，在腹部发育后产生临界浓度纳米蛋白，对于激活纳米的翻译至关重要。 纳米的定位是由一个复合物A在其3'未翻译区域（3'UTR）内的复合物A作用定位信号介导的。先前和初步研究的结果表明，细胞质定位因子在该定位信号中识别不同的序列或结构基序。拟议的工作将使用Nanos作为模型，将洞悉复杂的RNA定位信号如何通过细胞定位机械识别，以及这些RNA蛋白相互作用如何通过定位途径介导运输和锚定。更普遍地，这些研究将阐明RNA - 蛋白质相互作用提供对发展，生长和分化所需的基本细胞过程的高度选择性控制的机制。具体目标1包括对纳米定位信号的突变分析，以确定定位信号识别和功能的序列和结构要求。这项工作将通过从十种不同的果蝇种类的纳诺斯3'UTRS进行系统发育分析来促进这项工作。初步工作导致了一个候选纳米定位因子和第二秒的生化鉴定。 AIM 2着重于这些因素的生化和遗传表征，以确定其在纳米定位中的功能。此外，提出了一种新的生化分离策略。在AIM 3中，纳米定位因子的遗传筛选将补充AIM 2的生化方法。AIM4利用了一个新的系统用于体内Nanos RNA的GFP标记，以研究卵子发生过程中纳米定位的动态途径。