CYTOSKELETON-MEMBRANE INTERACTIONS

细胞骨架-膜相互作用

• 批准号: 3282378
• 负责人: ELIZABETH J. LUNA
• 金额: $ 15.83万
• 依托单位: WORCESTER FOUNDATION FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-07-01 至 1991-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282378
• 关键词: Dictyostelium; actins; affinity chromatography; antibody formation; binding proteins; cell adhesion; cell cell interaction; cell membrane; cell motility; chemotaxis; crosslink; cytoskeleton; density gradient ultracentrifugation; electron microscopy; enzyme linked immunosorbent assay; erythrocyte membrane; human tissue; immunoelectron microscopy; laboratory rabbit; membrane activity; membrane proteins; phagocytosis; protoplasm motility; radionuclides

A complete understanding of phenomena such as morphogenesis, wound healing, and viral transformation is contingent upon an understanding of fundamental cell processes such as motility, the maintenance and change of cell shape, and cell-cell and cell-substrate attachment. At the core of our ignorance about these basic life processes is a lack of knowledge about the molecular details of transmembrane interactions between the cytoskeleton and the cell surface. The ameboid stage of the cellular slime mold, Dictyostelium discoideum is easily grown in large quantities, is motile, and exhibits cell-cell as well as cell-substrate attachment. Using D. discoideum as a model for cells in general, I am investigating the interactions between this cell's plasma membrane and its cytoplasmic cytoskeletal proteins. In the past three years we have isolated cell-cell contact regions from aggregated amebae, and have developed two new low-speed sedimentation binding assays to monitor associations between actin and membranes. We find that integral proteins mediate a specific, saturable binding of plasma membranes to the sides of actin filaments and that plasma membranes can catalyze the polymerization of actin at the membrane surface even at actin concentrations well below the normal critical concentration for polymerization. Using detergent solubilization and F-actin affinity chromatography, we have identified an integral actin-binding protein with an apparent molecular weight of 17,000 daltons. The immediate goals are to characterize and reconstitute the 17kd protein, to use chemical crosslinking to identify detergent-sensitive actin-binding proteins missed by F-actin affinity chromatography, to prepare monospecific antibodies against the major integral membrane components of an ameboid membrane cytoskeleton, to explore the structure and regulation of the transmembrane linkages in this system, and to determine whether structurally similar linkages are found in other species. Of ultimate interest is the molecular basis for motility and cell-cell and cell-substrate interactions.

完全理解诸如形态发生，伤口愈合等现象， 病毒转化取决于对基本的理解 细胞过程，例如运动，维持和变化细胞形状， 以及细胞细胞和细胞基底附着。 我们无知的核心 关于这些基本的生活过程是缺乏有关分子的知识 细胞骨架与细胞之间的跨膜相互作用的细节 表面。 细胞粘液模具的肢体阶段，柱状 迪斯科医学很容易大量生长，易流动和展览 细胞细胞以及细胞基底附着。 将D. Discoideum用作 一般的细胞模型，我正在研究 该细胞的质膜及其细胞质细胞骨架蛋白。 在 在过去的三年中，我们从 汇总的Amebae，并开发了两个新的低速沉积物 结合测定法以监测肌动蛋白和膜之间的关联。 我们 发现整合蛋白介导了血浆的特定饱和结合 肌动蛋白丝的侧面，质膜可以 催化肌动蛋白在膜表面的聚合，即使在肌动蛋白处 远低于正常临界浓度的浓度 聚合。 使用洗涤剂溶解和F-肌动蛋白亲和力 色谱法，我们已经确定了一种整体肌动蛋白结合蛋白 明显的分子量为17,000个达尔顿。 直接目标是 特征并重建17KD蛋白，以使用化学 交联以识别缺失洗涤剂敏感肌动蛋白结合蛋白 通过F-肌动蛋白亲和色谱法，制备单特异性抗体 反对ameboid膜的主要整体膜成分 细胞骨架，探索跨膜的结构和调节 该系统中的链接，并确定结构相似 在其他物种中发现了联系。 最终感兴趣的是分子 运动性，细胞细胞和细胞基底相互作用的基础。