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（改编自申请人摘要）：细胞改变 形状是许多基本细胞过程的重要组成部分 包括运动、分裂和分化。 医学重要 形状变化包括血小板在血液凝固期间经历的形状变化 以及嗜中性粒细胞和巨噬细胞在吞噬作用期间所表现出的那些。的 这些形状变化的分子基础在于 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 这两种主要细胞骨架元素的组织状态 反过来又受到肌动蛋白结合和微管相关的宿主的调节 蛋白质（MAP）。 解释细胞形状变化的分子基础是 取决于对 ABP和MAP的体内功能。 一个优秀的模型实验系统 用于检查ABPs和MAPs功能的是海胆体腔细胞。 这些原始血细胞可以被认为是 脊椎动物的血小板和巨噬细胞，因为它们在 体腔液凝结，是高度吞噬性的，并且可以是能动的。 体腔细胞可以被诱导经历同步的，肌动蛋白介导的形状 从片状伪足到丝状伪足的变化以及相反的变化。 这一建议描述了一系列的形态和生化 旨在鉴定体腔细胞ABP和MAP的实验， 在形状变化过程中阐明它们的体内功能。 abp和 MAP将从肌动蛋白和微管蛋白结合的印迹覆盖物中鉴定。 细胞提取物，来自紫杉醇稳定的微管制剂和来自 DNA酶亲和层析。 光下免疫定位， 电子显微镜水平将被用来检查的分布 体腔细胞中的ABP和MAP，特别强调丝状伪足， 板状伪足形状改变。 这就允许检查 肌动蛋白细胞骨架的解体和MAP马达的功能 蛋白质在细胞器中的分布。作为次要目的， 钙调节体腔细胞的形状变化也将进行研究 使用使用钙离子载体处理和钙离子载体处理的实验， 依赖性荧光染料。 这些实验的结果应该 提供了新的见解的身份和功能的ABP和地图， 体腔细胞形状的变化，也应该有助于导致更好的 理解以下表现出的医学显著性形状变化 血小板和巨噬细胞。