MOLECULAR GENETIC ANALYSIS OF DEVELOPING SYNAPSES

 描述（由申请人提供）：神经细胞通信的主要方式是通过在化学突触处释放神经递质。大脑的功能以及大脑存储和处理信息的能力取决于这些精确可靠的突触连接。这项建议是针对发展一个分子的理解机制，调节这一过程的突触和突触的维护。我们建议使用遗传学，细胞生物学，分子生物学和实时成像的组合，以确定和表征的作用，通过研究一组神经-神经突触的特定组件的信号通路，协调突触发育的分子组成部分。在以前的工作中，我们描述了新生突触形成的细胞事件的顺序，通过可视化荧光标记的组件的招聘新形成的突触。此外，我们分离出了破坏这些突触形成的突变体。这些的一个子集的克隆确定了一个F-box蛋白，选择性靶向蛋白的泛素介导的降解，调节肌动蛋白和微管细胞骨架，转录调节蛋白和一种新的保守蛋白。使用各种分子，遗传和蛋白质相互作用的研究，我们现在建议，以确定在一个机制的水平，这些蛋白质如何发挥作用，以调节突触组装。此外，我们将使用遗传学的方法来分离和表征额外的基因，破坏机械感觉神经元和它们的突触伙伴之间的信号传导。elegans来扩展我们对这个过程的分子理解。这些方法将有助于定义细胞在突触形成和突触维持过程中用于识别和相互通信的机制。而C.与脊椎动物相比，已经很明显，这两个系统中存在类似的途径。因此，对参与C. elegans应该有助于定义一套一般的和可能保守的原则，这些原则通常是突触发生机制所共有的。

 DESCRIPTION (provided by applicant): The primary means by which nerve cells communicate is via the release of neurotransmitter at chemical synapses. The function of the brain and the ability of the brain to store and process information depend on these synaptic connections forming precisely and reliably. This proposal is directed towards developing a molecular understanding the mechanisms that regulated this process of synaptogenesis and synapse maintenance. We propose to use a combination of genetics, cell biology, molecular biology and live imaging to identify and characterize the role of molecular components of the signaling pathways that coordinate synaptic development by studying the specific assembly of a set of nerve-nerve synapses. In previous work, we described the order of cellular events in nascent synapse formation by visualizing the recruitment of fluorescent-tagged components to newly forming synapses. Furthermore, we isolated mutants that disrupt the formation of these synapses. Cloning of a subset of these identified an F-box protein that selectively targets proteins for ubiquitin-mediated degradation, regulators of both the actin and microtubule cytoskeleton, a transcriptional regulatory protein and a novel conserved protein. Using a variety of molecular, genetic and protein interaction studies we now propose to determine at a mechanistic level how these proteins function to regulate synapse assembly. In addition, we will use genetic approaches to isolate and characterize additional genes that disrupt signaling between mechanosensory neurons and their synaptic partners in C. elegans to extend our molecular understanding of the process. Together these approaches will help define mechanisms that cells use to identify and communicate with one another during the process of synapse formation and synaptic maintenance. While synaptogenesis is undoubtedly less complex in C. elegans than in vertebrates, it is already clear that similar pathways operate in both systems. Thus, analysis of the molecules participating in the process in C. elegans should help define a set of general and likely conserved principles that are common to synaptogenesis mechanisms in general.