RNA Localization in Xenopus Oocytes

描述（申请人提供）：细胞质RNA定位是产生胚胎和体细胞极性的广泛机制。在脊椎动物中，Vg1 mRNA是在胚胎模式中发挥作用的本地化mRNA的一个突出例子。Vg1 mRNA编码一种肽生长因子，并在卵发生过程中定位于爪蟾卵母细胞的植物细胞质中。卵细胞植物半球中Vg1蛋白的受限表达对于胚胎的正确模式至关重要，因此Vg1 mRNA的定位是理解母体分子如何定位以影响模式和极性的重要模型。本研究项目的目标是研究RNA分子如何靶向细胞质的特定区域，通过空间限制蛋白表达来产生细胞极性。我们对控制这一过程的分子相互作用的研究进展奠定了这项研究的基础。我们的实验已经定义了关键的rna -蛋白质和蛋白质-蛋白质相互作用，导致转运能力的RNP复合物的组装，并揭示了定位途径中的不同步骤。我们目前的建议是通过研究定位途径中这些步骤之间的动态转换来扩展这些发现。我们将结合体内和体外方法来确定Vg1 mRNA本地化表达的分子机制。提出了三个具体目标：在Aim I中，我们将描述细胞质Vg1 RNP复合物的动力学和运输特性，在Aim II中，我们将分析RNP运输颗粒的组装和活性，在Aim III中，我们将研究翻译抑制和RNA定位的耦合机制。提出的研究旨在揭示mRNA分子在细胞内运输以产生空间限制蛋白表达的机制。并将深入了解发育信号在脊椎动物胚胎中的空间分布。这项工作将影响与人类健康有关的问题，如出生缺陷，以及与RNA转运缺陷和局部蛋白质合成有关的神经系统疾病。本研究项目的目标是阐明负责RNA定位和局部蛋白合成的分子机制。这些过程对胚胎发育和神经功能都至关重要。特别是，某些人类疾病，包括脆性X智力迟钝综合征和脊髓性肌萎缩症，与RNA定位和局部蛋白合成缺陷有关。

DESCRIPTION (provided by applicant): Cytoplasmic RNA localization is a widespread mechanism for generating both embryonic and somatic cell polarity. Among vertebrates, Vg1 mRNA is a prominent example of a localized mRNA that plays a role in embryonic patterning. Vg1 mRNA encodes a peptide growth factor, and is localized during oogenesis to the vegetal cytoplasm of Xenopus oocytes. Restricted expression of Vg1 protein in the vegetal hemisphere of the egg is critical for correct patterning of the embryo, making localization of Vg1 mRNA an important model for understanding how maternal molecules are localized to influence pattern and polarity. The goal of this research project is to investigate how RNA molecules can be targeted to specific regions of the cell cytoplasm to generate cell polarity through spatially restricted protein expression. The foundation for this investigation has been laid by our progress studying the molecular interactions that govern this process. Our experiments have defined critical RNA-protein and protein-protein interactions leading to assembly of a transport-competent RNP complex and have uncovered distinct steps in the localization pathway. Our current proposal seeks to extend these findings by studying the dynamic transitions between these steps in the localization pathway. We will use a combination of in vivo and in vitro approaches to determine the molecular mechanisms responsible for localized expression of Vg1 mRNA. Three specific aims are proposed: In Aim I, we will characterize the dynamics and transport of the cytoplasmic Vg1 RNP complex, in Aim II, we will analyze the assembly and activity of RNP transport granules and in Aim III we will investigate the mechanisms that couple translational repression and RNA localization The proposed research is designed to reveal the mechanisms by which mRNA molecules are transported within cells to generate spatially restricted protein expression, and will provide insight into how developmental signals are spatially distributed in the vertebrate embryo. This work will impact issues related to human health such as birth defects, as well as neurological diseases that have been linked to defective RNA transport and localized protein synthesis. PUBLIC HEALTH RELEVANCE The goal of this research project is to elucidate the molecular mechanisms responsible for RNA localization and localized protein synthesis. These processes are critical for both embryonic development and neurological function. In particular, certain human diseases, including fragile X mental retardation syndrome and spinal muscular atrophy, have been linked to defects in RNA localization and localized protein synthesis.