ORGANELLE MOVEMENT IN FAST AXONAL TRANSPORT

说明神经元胞体是大多数轴突蛋白的位置。 综合。因此，维持和发展的必要材料 轴突必须沿着轴突从细胞体运输到 终点站。因此，轴突运输对神经生长和 再生和轴突运输缺陷导致神经变性 和疾病。本项目的目标是了解 轴突运输，细胞器及其货物的膜性有多大 在快速轴突运输中沿着轴突运输。基于两个微管的 蛋白质是这一运动的发动机。激动素是 细胞器顺行运输(从细胞体到细胞器 轴突终末)，胞浆动力蛋白逆行移动细胞器 方向，从末端到细胞体。然而，许多关于 顺行和逆行细胞器运动的协调仍然存在 被解释清楚。这项提议集中在逆行马达上， 胞质动力蛋白。人们已经发现，不同的动力蛋白复合体， 通过动力蛋白中间链的不同亚型来区分 亚单位(IC74)沿轴突顺行运输 在两个不同的隔间里。一个动力蛋白池在缓慢的轴突中移动 运输，并且第二池与膜细胞器相关联 被动蛋白所感动。这项提案的目的是为了理解该法规 以及这两个动力蛋白顺行库与 逆行动力蛋白马达。标记了IC74亚型的表位将是 将其导入培养细胞并鉴定其IC74亚型 与体内细胞器的逆行运动有关 下定决心。不同的动力蛋白复合体将被提纯，并 将确定每个复合体的功能特性，包括ATPase 活性、体外运动性和细胞器结合。中美之间的差异 确定了动力蛋白复合体的磷酸化状态。这个 动力蛋白的磷酸化状态将在体外被操纵，并发挥其作用 将确定在调节动力蛋白性质方面的磷酸化作用。

DESCRIPTION The neuronal cell body is the site of most axonal protein synthesis. Therefore, material necessary for the maintenance and growth of the axon must be transported from the cell body along the axon to the terminal. Thus, axonal transport is important for nerve growth and regeneration, and defects in axonal transport result in neurodegeneration and disease. The goal of this project is to understand one component of axonal transport, how membranous organelles, and their cargo, are transported along the axon in fast axonal transport. Two microtubule-based proteins are the motors for this movement. Kinesin is the motor for organelle transport in the anterograde direction (from the cell body to the axon terminal) while cytoplasmic dynein moves organelles in the retrograde direction, from the terminal to the cell body. However, many details about the coordination of anterograde and retrograde organelle movements remain to be explained. This proposal concentrates on the retrograde motor, cytoplasmic dynein. It has been found that distinct dynein complexes, which are distinguished by different isoforms of the dynein intermediate chain subunit (IC74), are transported along the axon in the anterograde direction in two different compartments. One pool of dynein is moving in slow axonal transport, and a second pool is associated with the membranous organelles moved by kinesin. The goal of this proposal is to understand the regulation of dynein and the relationship of these two anterograde pools of dynein to the retrograde dynein motor. Epitope tagged IC74 isoforms will be transfected into cultured cells and the identity of the IC74 isoforms which are associated with retrogradely moving organelles in vivo will be determined. The different dynein complexes will be purified and the functional properties of each complex will be determined, including ATPase activity, in vitro motility, and organelle binding. Differences in the phosphorylation state of the dynein complexes have been identified. The phosphorylation state of dynein will be manipulated in vitro and the role of phosphorylation in regulating the properties of dynein will be determined.