CYTOSKELETAL DYNAMICS DURING CELL SHAPE CHANGES

细胞形状变化期间的细胞骨架动力学

• 批准号: 2185133
• 负责人: JOHN H HENSON
• 金额: $ 9.72万
• 依托单位: DICKINSON COLLEGE
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-09-01 至 1995-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2185133
• 关键词: G protein; actin binding protein; affinity chromatography; blood cells; calcium flux; cell growth regulation; cell migration; cell morphology; cytoskeletal proteins; electron microscopy; fluorescent dye /probe; immunofluorescence technique; light microscopy; microtubule associated protein; organelles; phagocytosis; phase contrast microscopy; sea urchins; tubulin; video recording system

DESCRIPTION (Adapted from the applicant's abstract): Alterations of cell shape are an important component of many fundamental cell processes including movement, division, and differentiation. Medically important shape changes include those undergone by platelets during blood clotting and those exhibited by neutrophils and macrophages during phagocytosis. The molecular basis for these shape changes lies in the reorganization of cell's cytoskeletal proteins, chiefly the structural proteins actin and tubulin. The organization state of these two major cytoskeletal elements is in turn regulated by a host of actin binding and microtubule associated proteins (MAP). Explaining the molecular basis of cell shape changes is dependent on the understanding of the in vivo function of ABPs and MAPs. One excellent model experimental system for examining the functions of ABPs and MAPs is the sea urchin coelomocyte. These primitive blood cells can be considered as a cross between the vertebrate platelet and macrophage in that they change shape during coelomic fluid coagulation, are highly phagocytic and can be motile. Coelomocytes can be induced to undergo synchronous, actin mediated shape changes from a lamellipodial to a filopodial form as well as the reverse. This proposal describes a series of morphological and biochemical experiments aimed at the identification of coelomocyte ABPs and MAPs and the elucidation of their in vivo functions during shape changes. ABPs and MAPs will be identified from actin and tubulin binding overlays of blots of cell extracts, from taxol stabilized microtubule preparations and from DNase affinity chromatography. Immunolocalization at the light and electron microscopic levels will be used to examine the distribution of ABPs and MAPs in coelomocytes with particular emphasis on the filopodial to lamellipodial shape change. This allows for the examination of the dismantling of the actin cytoskeleton and the function of MAP motor proteins in the distribution of cellular organelles.As a secondary aim, the calcium regulation of coelomocyte shape changes will also be investigated using experiments employing calcium ionophore treatments and calcium dependent fluorescent dyes. The results of these experiments should provide new insights into the identity and functions of ABPs and MAPs in coelomocyte shape changes and should also help lead to a better understanding of the medically significant shape changes exhibited by platelets and macrophages.

描述（改编自申请人的摘要）：细胞的更改 形状是许多基本细胞过程的重要组成部分 包括运动，分裂和差异化。 重要的是重要的 形状的变化包括血液凝结过程中血小板经历的变化 以及吞噬作用期间中性粒细胞和巨噬细胞表现出的。这 这些形状变化的分子基础在于重组 细胞的细胞骨架蛋白，主要是结构蛋白肌动蛋白和 微管蛋白。 这两个主要细胞骨架元素的组织状态 反过 蛋白质（地图）。 解释细胞形状变化的分子基础是 取决于对 ABP和地图的体内功能。 一个出色的模型实验系统 为了检查ABP和地图的功能，是海胆核细胞。 这些原始的血细胞可以视为 脊椎动物的血小板和巨噬细胞，因为它们会改变形状 腔液凝结，高度吞噬，可能是流动性的。 可以诱导核细胞进行同步，肌动蛋白介导的形状 从层状形式变为丝状形式以及反向。 该建议描述了一系列形态学和生化 旨在鉴定核细胞ABP和地图的实验，以及 在形状变化过程中，阐明其体内功能。 ABP和 地图将从肌动蛋白和微管蛋白结合覆盖层的印迹中识别 细胞提取物，来自紫杉醇稳定的微管制剂，并从 DNase亲和力色谱法。 在光线下进行免疫定位 电子显微镜水平将用于检查 卵形细胞中的ABP和地图特别强调了丝状 层状形状变化。 这允许检查 拆卸肌动蛋白细胞骨架和地图电机的功能 细胞器分布中的蛋白质。作为次要目的， 还将研究核细胞形状变化的钙调节 使用采用钙离子载体治疗和钙的实验 依赖性荧光染料。 这些实验的结果应 提供有关ABP和地图的身份和功能的新见解 核细胞形状的变化，也应有助于导致更好 了解医学上显着的形状变化所表现出的 血小板和巨噬细胞。