PLASMA MEMBRANE REGIONS AS CELLS GROW IN MONOLAYERS

细胞单层生长时的质膜区域

• 批准号: 3276619
• 负责人: BRUCE Shell JACOBSON
• 金额: $ 13.7万
• 依托单位: UNIVERSITY OF MASSACHUSETTS AMHERST
• 依托单位国家: 美国
• 财政年份: 1981
• 资助国家: 美国
• 起止时间: 1981-04-01 至 1992-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3276619
• 关键词: HeLa cells; affinity chromatography; apical membrane; basement membrane; basolateral membrane; cell adhesion; cell cell interaction; cell membrane; crosslink; cytoskeleton; electron microscopy; extracellular matrix; gel electrophoresis; human tissue; laboratory mouse; lipid biosynthesis; membrane activity; membrane proteins; membrane structure; monoclonal antibody; protein biosynthesis; radiotracer; tissue /cell culture

The long term objective of the proposed research is to understand the role of the plasma membrane (PM) proteins and the cytoskeleton in mediating the formation of apical and basolateral domains of the plasma membrane in cells that exhibit a transcellular polarity. Examples of such cells are epithelial cells which are bounded on one side by a fluid phase and on the other by an extracellular matrix. In vitro adhesion of HeLa cells (a transformed epithelium easily grown in suspension) to an extracellular matrix protein, gelatin, will be used as a model to study the formation of the apical and basolateral PM. In this system the initial stages in the formation of these PM domains are cell attachment and spreading. HeLa cells attached to gelatin coated culture dishes are induced to spread rapidly and form an apical PM exposed to the culture medium and a basal PM adjacent to the culture dish. The apical and basal domains can be isolated and redistribution of proteins between the domains during spreading can be quantitated. We have identified five gelatin receptors on the HeLa cell surface. The kinetics of cell attachment and spreading indicate that spreading is a cooperative process leading to the hypothesis that the mechanism for spreading is the segregation of the cell surface extracellular matrix receptors into the basal PM with subsequent clustering of the receptors into oligomers inducing them to bind to the cytoskeleton. The hypothesis will be tested by concentrating on several specific aims: 1, determine which of the known gelatin receptors mediate attachment and/or spreading; 2, determine which receptors redistribute and become segregated among the apical, basal and internal PM domains during cell adhesion; 3, determine if gelatin receptor clustering is correlated with cell spreading; 4, determine if receptor binding to the cytoskeleton is regulated by receptor clustering and/or covalent modification; 5, determine which cytoskeletal proteins bind to the gelatin receptors and; 6, determine if the receptors are transmembrane proteins present in oligomeric complexes and if they have common polypeptide fragments. Results from the specific aims will undoubtedly increase our understanding of the molecular mechanism of cell adhesion to an extracellular matrix and the formation of transcellular polarity.

拟议研究的长期目标是了解 质膜（PM）蛋白和 介导顶角和基底外侧的形成中的细胞骨架 质膜的结构域在细胞中表现出A 细胞极性。 这种细胞的例子是上皮细胞 由流体阶段在一侧的边界，另一侧由 细胞外基质。 HeLa细胞的体外粘附（A 在悬浮液中变化的上皮） 细胞外基质蛋白明胶，将用作模型 研究顶端和基底外侧PM的形成。 在这个 系统这些PM域的形成的初始阶段 是细胞的附着和扩散。 HELA细胞附着在明胶上 涂层培养皿被诱导迅速扩散并形成 顶端PM暴露于培养基和基础PM附近 到文化菜。 可以隔离顶端和基础域 和在域之间的蛋白质重新分布 可以定量扩展。 我们已经确定了五种明胶 HELA细胞表面上的受体。 细胞动力学 依恋和传播表明传播是合作社 过程导致假设是 扩散是细胞表面细胞表面的分离 基础PM的基质受体，随后的聚类 受体进入低聚物，诱导它们与 细胞骨架。 该假设将通过集中于 几个具体目的：1，确定哪些已知明胶 受体介导依恋和/或扩散； 2，确定 哪个受体重新分布并在 细胞粘附过程中的顶，基础和内部PM结构域； 3， 确定明胶受体簇是否与细胞相关 扩散； 4，确定受体与细胞骨架的结合是否为 由受体聚类和/或共价修饰调节； 5， 确定哪种细胞骨架蛋白与明胶结合 受体和； 6，确定受体是否是跨膜 寡聚复合物中存在的蛋白质，如果它们具有 常见的多肽片段。 特定目的的结果 无疑会增加我们对分子的理解 细胞粘附对细胞外基质的机理和 跨细胞极性的形成。