REGULATION OF NEURONAL ACTIN BY BETA-THYMOSINS

肌动蛋白是神经细胞中一种主要的细胞骨架蛋白，参与 在生长锥运动、轴突等多种运动功能中 运输和神经递质释放。在细胞内，肌动蛋白单体和 聚合物以动态平衡的形式存在，这取决于 控制肌动蛋白速率的肌动蛋白结合蛋白之间的相互作用 成核、肌动蛋白单体的可用性以及长度和数量 细丝的。每类调控蛋白都已在 大脑。最近，胸腺肽Beta4(Tbeta4)，一种被认为是 在大脑中大量表达，已被确定为肌动蛋白 单体结合蛋白，可能在维持 很大比例的肌动蛋白是未聚合的。它抑制了 肌动蛋白自发成核，但允许肌动蛋白从未封顶的 肌动蛋白核团。这提供了一种简单的空间协调机制 以及对细胞内肌动蛋白聚合级联的时间控制。在……里面 在这笔赠款中，我们将研究β-胸腺素在调节 神经元中肌动蛋白的组装和拆解。因为β-胸腺素 功能必须在多组分系统的环境中考虑， 我们还将描述肌动蛋白明胶蛋白的特征，但不太详细。 切断和封端结合肌动蛋白单体的蛋白质和Profilin 蛋白。首先，我们将确定Tbeta4是否调节生长锥 运动性和肌动蛋白动力学。Tbeta4在神经元中的分布将 接受检查。Tbeta4将被显微注射到神经元中，并对 将监测生长锥运动和肌动蛋白动态。第二，到 检验神经诱导PC12细胞中的Tbeta4的假设 生长因子通过诱导肌动蛋白促进轴突延伸 解聚。批量加载将提高PC12中的Tbeta4水平 或表达载体的转染法。Tbeta4水平将会降低 批量负载抗Tbeta4抗体并与反义核酸孵育 核苷酸。第三，确定Tbeta4对调控是否重要 突触体和生长锥标本中肌动蛋白动力学的研究。第四, 确定Tbeta4肌动蛋白结合位点，它也可能存在于 其他几种肌动蛋白结合蛋白。结合使用显微技术， 生化和分子生物学方法将提供更好的 了解肌动蛋白细胞骨架是如何在几个 神经元的子域。显微、生化的结合使用 分子生物学方法将提供更好的理解 肌动蛋白细胞骨架是如何在不同的亚区维持的 神经元。

Actin is a major cytoskeletal protein in neuronal cells, and is involved in a variety of motile functions such as growth cone motility, axonal transport and neurotransmitter release. In the cell, actin monomers and polymers exist in a dynamic equilibrium which is dependent on the interplay between actin binding proteins controlling the rate of actin nucleation, the availability of actin monomers and the length and number of filaments. Each class of regulatory proteins have been identified in the brain. Recently, thymosin beta4 (Tbeta4), a protein which is abundantly expressed in the brain, has been identified as an actin monomer binding protein which may play a significant role in maintaining a large proportion of the actin in the unpolymerized form. It suppresses spontaneous actin nucleation but allows actin to polymerize from uncapped actin nuclei. This provides a simple mechanism for coordinating spatial and temporal control of the actin polymerization cascade in cells. In this grant, we will examine the role of beta-thymosins in regulating actin assembly and disassembly in neurons. Because beta-thymosin function must be considered in the context of a multi-component system, we will also characterize, but in less detail, gelsolin, an actin severing and capping protein and profilin, an actin monomer binding protein. First, we will determine if Tbeta4 regulates growth cone motility and actin dynamics. The distribution of Tbeta4 in neurons will be examined. Tbeta4 will be microinjected into neurons and effects on growth cone motility and actin dynamics will be monitored. Second, to test the hypothesis that Tbeta4, which is induced in PC12 cells by nerve growth factor, promotes neurite extension by inducing actin depolymerization. Tbeta4 level in PC12 will be raised by bulk loading or transfection of an expression vector. Tbeta4 level will be decreased by bulk loading of anti-Tbeta4 antibody and incubation with antisense nucleotides. Third, determine if Tbeta4 is important for the regulation of actin dynamics in synaptosome and growth cone preparations. Fourth, identify the Tbeta4 actin binding site, which may also be present in several other actin binding proteins. The combined use of microscopic, biochemical and molecular biological approaches will provide a better understanding of how the actin cytoskeleton is maintained in several subdomains of the neuron. The combined use of microscopic, biochemical and molecular biological approaches will provide a better understanding of how the actin cytoskeleton is maintained in different subdomains of the neuron.