Mechanism and Regulation of Actin-Based Retrograde Flow

描述（由申请人提供）：逆行流动是一种由肌动蛋白细胞骨架介导的细胞运动形式，广泛存在于真核细胞中。流动过程涉及板状脂质体质膜、膜蛋白和下面的肌动蛋白细胞骨架从细胞外围向细胞中心的持续运动。这种现象已被表明是重要的细胞易位，在靶向细胞迁移，在细胞基质相互作用，并在调节肌动蛋白和微管之间的串扰。逆行流动已被广泛研究，只有少数细胞类型（特别是神经元和哺乳动物和两栖动物组织培养细胞），尽管这些努力，确切的机制和调节这一基本过程仍然是未知的。这项建议的目的是扩大知识的机制背后的逆行流动使用一个独特的实验模型，海胆体腔细胞。这些细胞显示出一种高度夸张的逆行流动形式，并具有许多属性，使它们非常适合本研究，包括它们的光学性质，细胞骨架蛋白的免疫探针的可用性，以及流动可以开始和停止以及细胞骨架组织可以改变的准备。光学和电子显微镜方法，结合药理学和显微操作方法，将用于解决以下特定目的： 1.确定体腔细胞逆行流动中肌动蛋白丝与肌动蛋白结合蛋白和运动蛋白之间的结构和功能关系 2.阐明体腔细胞逆行流动中的物理作用力。 3.分析体腔细胞逆向流动中肌动蛋白丝和微管之间的结构和功能关系。 4.目的：研究体腔细胞由板状伪足向丝状伪足的诱导转化。

DESCRIPTION (provided by applicant): Retrograde flow is a form of cell motility that is mediated by the actin cytoskeleton and is widespread in eukaryotic cells. The flow process involves the continual movement of the lamellipodial plasma membrane, membrane proteins and underlying actin cytoskeleton from the cell periphery towards the cell center. This phenomenon has been indicated as being important in cellular translocation, in the targeting of cellular migrations, in cell-substrate interactions, and in modulating the crosstalk between actin and microtubules. Retrograde flow has been extensively studied in only a few cell types (particularly neurons and mammalian and amphibian tissue culture cells) and, despite these efforts, the exact mechanism and regulation of this fundamental process is still largely unknown. The objective of this proposal is to expand the knowledge of the mechanisms underlying retrograde flow using a unique experimental model, the sea urchin coelomocyte. These cells display a highly exaggerated form of retrograde flow and possess a number of attributes which make them well suited for this study, including their optical properties, the availability of immunological probes for cytoskeletal proteins, and the readiness with which flow can be started and stopped and the cytoskeletal organization can be altered. Light and electron microscopic methods, combined with pharmacological and micromanipulation approaches will be used to address the following Specific Aims: 1. To determine the structural and functional relationships between actin filaments and actin-binding and motor proteins in the context of coelomocyte retrograde flow 2. To elucidate the physical forces operating in the context of coelomocyte retrograde flow. 3. To analyze the structural and functional relationships between actin filaments and microtubules in the context of coelomocyte retrograde flow. 4. To study the inducible transformation of coelomocytes from a lamellipodial to a filopodial form.