Collaboration between actin nucleators - Spire and Cappuccino

项目摘要 细胞骨架对生命的许多方面都是必不可少的，细胞分裂和运动是主要的例子。的 该建议的首要目标是了解两种肌动蛋白成核剂Spire（Spir）和 卡布奇诺（卡普）。我们正在研究果蝇中的这些蛋白质，在果蝇中， 极性在早期发育中建立。Spir和Capu构建了一个actin网格， 果蝇卵母细胞在整个中期卵子发生。补片的存在及其在正确位置的移除 发育阶段对极性建立都是至关重要的。从而了解Spir和Capu如何合作， 建立一个网状结构对于基本理解它们如何对极性的建立做出贡献是至关重要的 在发展过程中。在建议的工作中，我们将重点关注Spir-Capu操作的两个方面：1） 机制，他们组装肌动蛋白丝和2）如何Spir和Capu是时空 调节，以建立肌动蛋白网，然后在适当的时候将其移除。我们已经开发出一种 Spir和Capu在整个卵子发生中期（阶段5-9）都很活跃的工作模型; Spir增强了 成核由Capu，特别是在膜;他们一起建立一个动态肌动蛋白网格，直到阶段10时， Spir和Capu蛋白质水平下降，网格消失，开始流动。我们将测试这个模型， 在体外和体内方法中， 从显微镜到整个动物遗传学测试这个模型将导致对两个问题的机械理解 蛋白质，保守的整个后生动物物种。它将促进我们对细胞骨架的了解， 它是受控的。考虑到这对蛋白质在极性细胞（包括神经元和上皮细胞）中的共存， 在哺乳动物细胞中，已经提出它们作为极性因子在果蝇中的作用是保守的。因此我们 我希望我们在果蝇卵子发生中对Spir和Capu的了解将适用于我们的研究。 了解许多动物的细胞骨架，细胞极性，生育力，发育和健康， 从果蝇到人类

Project Summary The cytoskeleton is essential to many aspects of life, cell division and motility being prime examples. The overarching goal of this proposal is to understand the roles of two actin nucleators, Spire (Spir) and Cappuccino (Capu). We are studying these protein in Drosophila, where they are known to be essential to polarity establishment during early development. Spir and Capu build an actin mesh that traverses the Drosophila oocyte throughout mid-oogenesis. The presence of the mesh and its removal at the correct developmental stage are both critical to polarity establishment. Thus learning how Spir and Capu collaborate to build the mesh is essential to a fundamental understanding of how they contribute to polarity establishment during development. In the proposed work, we will focus on two aspects of Spir-Capu operation: 1) the mechanism by which they assemble actin filaments and 2) how Spir and Capu are spatially and temporally regulated, in order to establish the actin mesh and then remove it when appropriate. We have developed a working model in which Spir and Capu are both active throughout mid-oogenesis (stages 5-9); Spir enhances nucleation by Capu, specifically at membranes; together they build a dynamic actin mesh until stage 10 when Spir and Capu protein levels drop, the mesh disappears, and streaming begins. We will test this model using both in vitro and in vivo approaches with assays that span scales from single molecule fluorescence microscopy to whole animal genetics. Testing this model will lead to a mechanistic understanding of two proteins, conserved throughout metazoan species. It will advance our knowledge of the cytoskeleton and how it is controlled. Given the co-existence of this pair of proteins in polar cells, including neurons and epithelial cells in mammals, it has been proposed that their role as polarity factors in Drosophila is conserved. Thus we anticipate that what is learned about Spir and Capu in Drosophila oogenesis will be applicable to our understanding of the cytoskeleton, cell polarity, fertility, development and health in many animals, ranging from Drosophila to humans.