ASPECTS OF CADHERIN/CATENIN COMPLEXES

钙粘蛋白/连环蛋白复合物的方面

• 批准号: 6386046
• 负责人: MARGARET J WHEELOCK
• 金额: $ 30.74万
• 依托单位: UNIVERSITY OF NEBRASKA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-08-01 至 2002-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6386046
• 关键词: animal tissue; cadherins; cell adhesion; cell cell interaction; cell motility; cell transformation; fibroblast growth factor; growth factor receptors; hepatocyte growth factor; human tissue; immunofluorescence technique; immunoprecipitation; laboratory mouse; polymerase chain reaction; protein binding; protein protein interaction; protein structure function; tissue /cell culture; western blottings; yeast two hybrid system

The long-term goal of this project is to understand the role cadherins and catenins play in cell-cell adhesion, both as structural components of the adherens junction and as cellular signaling molecules. The components of he adherens junction have been shown to play an important role in maintenance of normal tissues. Mutations in the proteins that make up these junctions often result in tumor formation. In some cases, this is due to inability of the cells to form the physical structure, that is, the junction and in other cases it is due to alterations in signaling pathways. The cadherin, as the transmembrane components of the adherens junctions, have been linked to the wnt or wingless signaling pathway. The wnt pathway also includes APC which is often mutated in carcinomas, particularly in colon carcinomas. A complete picture of the protein- protein interactions involved in the components of this pathway will lead to a better understanding of how cells become tumorigenic and eventually to novel diagnostics and/or treatments for carcinomas. The specific aims of this proposal are focused on understanding some of the properties of cells that are changes during tumor formation and that are dependent upon the cadherin/catenin complex. Specifically, we propose to: 1. Establish a role for cadherins in cell motility. 2. Characterize structural and functional differences between beta-catenin and plakoglobin (gamma-catenin). 3. Examine alpha-catenin for additional binding proteins. To accomplish these goals, we will make use of molecular constructs, often made up of chimeric molecules. These constructs will be transfected into mammalian tissue culture cells to observe their effect on cellular behavior and phenotype. Protein-protein interactions will be determined by several means which include co-immunoprecipitation reactions, co- localization in tissue culture cells and yeast two hybrid analysis. Protein-protein interactions will also be analyzed by co-transfecting interaction domains of two proteins into eukaryotic cells followed by co- localization in intact cells (immunofluorescence) and co- immunoprecipitation of cell extracts.

这个项目的长期目标是了解钙粘蛋白的作用， 连环蛋白在细胞-细胞粘附中起作用，两者都是细胞粘附的结构组分。 粘附连接和作为细胞信号分子。的组件 粘附连接已显示在以下方面起重要作用 维持正常组织。组成这些蛋白质的突变 这些连接经常导致肿瘤形成。在某些情况下，这是 由于细胞不能形成物理结构，即， 在其他情况下，这是由于信号传导的改变， 途径。钙粘蛋白作为粘附分子的跨膜成分， 连接，已经被连接到wnt或无翅信号通路。的 wnt途径还包括APC，其通常在癌中突变， 特别是在结肠癌中。蛋白质的完整图片- 蛋白质相互作用参与这一途径的组成部分将导致 更好地理解细胞是如何成为致瘤性的， 涉及癌的新诊断和/或治疗。 本提案的具体目标是着重了解以下一些问题： 细胞的特性在肿瘤形成过程中发生变化， 依赖于钙粘蛋白/连环蛋白复合体。具体来说，我们建议 收件人： 1.建立钙粘蛋白在细胞运动中的作用。 2.描述β-连环蛋白之间的结构和功能差异 和斑珠蛋白（γ-连环蛋白）。 3.检查α-连环蛋白是否有其他结合蛋白。 为了实现这些目标，我们将利用分子构建体， 由嵌合分子组成。这些构建体将被转染到 哺乳动物组织培养细胞，观察其对细胞增殖的影响。 行为和表型。蛋白质-蛋白质相互作用将通过以下方式确定： 几种方法包括免疫共沉淀反应，共- 组织培养细胞定位和酵母双杂交分析。 蛋白质-蛋白质相互作用也将通过共电泳分析 两种蛋白质的相互作用结构域进入真核细胞， 在完整细胞中的定位（免疫荧光）和共 细胞提取物的免疫沉淀。