STRUCTURE AND FUNCTION OF LENS COMMUNICATING CHANNELS

镜头通信通道的结构和功能

• 批准号: 3258584
• 负责人: GUIDO A ZAMPIGHI
• 金额: $ 11.37万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 1982
• 资助国家: 美国
• 起止时间: 1982-08-01 至 1993-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3258584
• 关键词: axon; biological information processing; calmodulin; electron microscopy; gap junctions; image processing; immunocytochemistry; laboratory rat; lens; lens proteins; liver cells; membrane permeability; membrane reconstitution /synthesis

The ultimate purpose of the research proposed in this application is to understand the structural organization and mechanisms of action of gap junctions. These organelles are thought to mediate and regulate the passage of low molecular weight molecules and ions between adjacent cells in an enormous variety of animal tissues. The unraveling of the mechanisms involved in cell-cell communication will be fundamental to further our understanding of normal tissue physiology (i.e., cardiac muscle), development and cell differentiation, and pathological processes such as cancer and lens cataracts. Membranes isolated from lenses of calves and mature bovines contain junctions of different morphology (16-18nm thick, 12-14nm thick, and wavy) and protein composition. They are being studied by electron microscopy, immunocytochemistry, and biochemical and electrical methodology. Specifically, the studies proposed will focus on the localization and electrical characteristics of two putative junction-forming proteins of the lens: MIP26 (the main intrinsic protein of the lens) and MP70 (present in the cortex of lenses from mature animals and in both cortex and nucleus of lenses of calves). The electrical properties of MIP26 and MP70 can be studied independently since these proteins can be isolated separately by using non-ionic detergents at low concentrations. The proteins are then completely solubilized with high concentration of the non-ionic detergent octylglucoside, reconstituted into single-walled liposomes, and fused with planar lipid films. The single and multi-channel properties of these proteins are characterized and compared with each other and those of liver gap junction proteins reconstituted by others. This application also proposes to study the molecular architecture of the gap junction channel at the secondary structural level. Hydrophobicity plots of the primary structure of liver hepatocytes and heart myocite gap junction protein strongly suggest that each monomer is constructed of four intramembrane spanning domains. Similar plots calculated from the primary structure of lens MIP26 suggest six membrane spanning domains. Cryo-electron microscopy and computer image processing will be utilized to experimentally test these predictions.

本应用程序提出的研究的最终目的 是了解结构组织和机制 间隙连接的动作。 这些细胞器被认为是介导的 并调节低分子量分子和 众多动物中相邻细胞之间的离子 组织。 涉及细胞细胞的机制的解开 沟通将是我们对我们对的理解至关重要的 正常组织生理（即心肌），发育和 细胞分化和病理过程，例如癌症 和镜头白内障。 从小牛和成熟牛透镜中分离出的膜 包含不同形态的连接（16-18nm厚，12-14nm） 厚，波浪形）和蛋白质组成。 他们正在研究 通过电子显微镜，免疫细胞化学和生化和 电气方法。 具体而言，提出的研究将 专注于两个的本地化和电特性 镜头的假定结形成蛋白：MIP26（主要 镜头的内在蛋白质）和 MP70（存在于成熟动物的镜片皮层中 犊牛镜头的皮质和核）。 电气 可以独立研究MIP26和MP70的属性 这些蛋白质可以通过非离子化分别分离 低浓度的洗涤剂。 蛋白质是 完全用高浓度的非离子化溶解 清洁剂八糖苷，重构为单壁 脂质体，并与平面脂质膜融合。 单曲和 这些蛋白质的多通道特性是表征的，并且 彼此和肝间隙连接蛋白的相比 由他人重组。 该应用还建议研究分子 间隔频道的架构 结构水平。 主要结构的疏水图 肝肝细胞和心肌间隙连接蛋白 强烈建议每个单体由四个 膜内跨越域。 从 镜头MIP26的主要结构表明六个膜跨度 域。 冷冻电子显微镜和计算机图像处理 将用于实验测试这些预测。