Protein Kinase C-mediated Integrin Activation

蛋白激酶 C 介导的整合素激活

• 批准号: 6525729
• 负责人: Jianxun LI
• 金额: $ 23.38万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 1996
• 资助国家: 美国
• 起止时间: 1996-08-01 至 2005-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6525729
• 关键词: actin binding protein; biological signal transduction; calmodulin; cell membrane; cytoskeletal proteins; cytoskeleton; enzyme substrate complex; fluorescence resonance energy transfer; integrins; intermolecular interaction; leukocyte adhesion molecules; leukocytes; membrane proteins; microtubule associated protein; molecular assembly /self assembly; phosphorylation; protein binding; protein degradation; protein kinase C; protein localization; protein purification; protein structure function; site directed mutagenesis; tissue /cell culture

DESCRIPTION (provided by applicant): In resting leukocytes, B2 integrin molecules are immobilized on the cell membrane in a diffused distribution, because they are constrained by cytoskeletal complex. Activation signals through the "inside-out" pathway cause this cytoskeletal complex to dissociate, resulting in enhanced free motion and clustering of f32 integrin. The clustered integrin provides high avidity binding to the ligands. The goal of this proposal is to identify and characterize the cytoskeletal complex constraining the integrin molecules in resting leukocytes, termed "preactivation cytoskeletal complex" in this proposal. Using the ability of regulating molecular mobility and clustering of B2 integrin as the criteria, we propose to determine whether talin, calmodulin, and dynamitin - three proteins involved in integrin activation-are members of the preactivation complex. Since integrin activation should not be a "none" or "all" process, two quantitative assays will be introduced: the single particle tracking method to quantitate the molecular mobility of B2 integrin and the fluorescent resonance energy transfer method to quantitate the clustering. Once we have identified the new members of the preactivation complex, their interrelationship with the currently known members, i.e. PKC, MacMARCKS, actin, and microtubule filaments, and among themselves will be determined by individually disrupting the function of each one by mutagenesis or by specific inhibitors. The significance of these studies is two-fold: if these proteins are indeed members of the preactivation complex, then the currently understood sequence of integrin activation is confirmed and we will have found new members. On the other hand, if these proteins regulate integrin activation without changing integrin mobility and clustering, then the currently held scheme of integrin activation requires modification. Information obtained from this study will provide a new understanding of the role of cytoskeletal proteins in regulating integrin activation. Since B2 integrin is a crucial regulator of the immune function in health and in diseases, this proposal has broad significance for clinical medicine.

性状（由申请方提供）：在静息白细胞中，B2整联蛋白 分子以扩散分布固定在细胞膜上， 因为它们受到细胞骨架复合体的限制。激活信号 通过"由内向外"途径导致这种细胞骨架复合物解离， 导致F32整联蛋白的自由运动和聚集增强。群集 整联蛋白提供与配体的高亲合力结合。这个目标 建议是确定和表征细胞骨架复合物约束 静息白细胞中的整合素分子，称为"预激活 细胞骨架复合物”在这一建议。 利用B2调节分子运动和聚集的能力， 整合素作为标准，我们建议确定是否talin，钙调素， 和dynamitin--三种参与整合素激活的蛋白质--是 激活前复合体由于整合素激活不应该是"无"或"无"， 在“全”过程中，将引入两种定量分析：单颗粒 跟踪方法定量B2整联蛋白的分子迁移率， 荧光共振能量转移法定量聚簇。一旦 我们已经确定了前激活复合体的新成员， 与目前已知的成员，即PKC，MacMARCKS，肌动蛋白， 和微管细丝，它们之间将由 通过诱变或通过特异性的 抑制剂的 这些研究的意义是双重的：如果这些蛋白质确实是 成员的前激活复合物，那么目前所理解的序列 整联蛋白激活得到确认，我们将发现新的成员。上 另一方面，如果这些蛋白质调节整合素活化而不改变 整合素的移动性和聚集性，那么目前所持有的整合素的方案 激活需要修改。 从这项研究中获得的信息将提供一个新的理解， 细胞骨架蛋白在调节整合素活化中的作用。自B2以来 整合素是健康和免疫系统中免疫功能的重要调节因子。 这一建议对临床医学具有广泛的意义。