BIOCHEMISTRY OF THE NEURONAL CYTOSKELETON

本提案的总体目标是了解结构， 神经元细胞骨架的功能。 拟议的研究旨在 让我们对神经细胞骨架的功能有一个基本的了解 在正常的中枢神经系统。 为此，我们将使用各种 技术来研究表达，组装，磷酸化和 许多神经元细胞骨架蛋白的相互作用。 最终我们 我希望这些信息能帮助我们理解为什么以及如何神经元 细胞骨架受到某些神经系统疾病的干扰，例如 阿尔茨海默氏病和肌萎缩侧索硬化症以及暴露于 神经毒素，如铝和丙烯酰胺。 本建议的具体目标是：（1）研究相互作用， 使用纯化的酶和蛋白质的三种神经丝蛋白的组装 每个神经丝三联体分子的溴化氰衍生片段 为了确定这三种蛋白质的磷酸化作用， 神经丝组装; 2）鉴定和分离激酶和/或 负责神经丝磷酸化的磷酸酶 蛋白质; 3）继续我们对神经丝相互作用的研究 和微管，以确定哪些微管相关蛋白 （地图）参与这些相互作用; 4）研究的重要性， 细胞骨架相关的ATP酶，β-interexin 神经元和培养细胞中的胞质转运 和5）获得高分子量神经丝的cDNA探针 蛋白质，NF-M和NF-H，将用于研究表达和 这些蛋白质的功能，并研究各种修饰的组装 proteins. 将cDNA克隆置于表达载体中，并将其 分布将在细胞中进行研究，这些细胞通常不包含 神经丝 这些研究将使我们能够确定 它们在细胞中的形成以及它们的功能。

The overall goal of this proposal is to understand the structure and function of the neuronal cytoskeleton. The proposed studies are designed to give us a basic knowledge of the function of the neuronal cytoskeleton in the normal CNS. Towards this end, we will use a wide variety of techniques to study the expression, assembly, phosphorylation and interactions of a number of neuronal cytoskeletal proteins. Ultimately, we hope that this information will help us understand why and how the neuronal cytoskeleton is perturbed by certain neurological diseases, such as Alzheimer's disease and amyotropic lateral sclerosis and by exposure to neurotoxins, such as aluminum and acrylamide. The specific aims of this proposal are 1) to study the interactions and assembly of the three neurofilament proteins using purified enzymatic and cyanogen bromide derived fragments of each neurofilament triplet molecule to determine the role of phosphorylation of these three proteins in the neurofilament assembly; 2) to identify and isolate the kinase and/or phosphatase responsible for the phosphorylation of the neurofilament proteins; 3) to continue our studies on the interactions of neurofilaments and microtubules, in order to define which microtubule associated proteins (MAPs) are involved in these interactions; 4) to study the importance of the cytoskeletal associated ATPase, Beta-internexin in cytoskeleton-mediated cytoplasmic transport in neurons and cultured cells and 5) to obtain cDNA probes for the high molecular weight neurofilament proteins, NF-M and NF-H, which will be used to study the expression and function of these proteins and to study the assembly of various modified proteins. The cDNA clones will be placed in expression vectors and their distribution will be studied in cells, which do not normally contain neurofilaments. These studies will allow us to determine what structures they form in the cell and what their function might be.