ASPECTS OF CADHERIN/CATENIN COMPLEXES

这个项目的长期目标是了解钙粘素和钙粘附素的作用 连环蛋白在细胞间的黏附中发挥作用，两者都是 粘着连接和作为细胞信号分子。的组件 他的黏附连接已被证明在 对正常组织的维护。构成蛋白质的突变 这些连接常常会导致肿瘤的形成。在某些情况下，这是 由于细胞不能形成物理结构，即 连接，在其他情况下是由于信令的改变 小路。作为粘附剂跨膜成分的钙粘附素 连接，已经连接到WNT或无翼信号通路。这个 WNT途径还包括在癌症中经常突变的APC， 尤其是在结肠癌中。一张完整的蛋白质图片- 参与这一途径各组成部分的蛋白质相互作用将导致 为了更好地理解细胞如何变得致瘤并最终 涉及癌症的新诊断和/或治疗方法。 本提案的具体目标集中在理解以下几个方面 在肿瘤形成过程中发生变化的细胞的特性 依赖于钙粘附素/连环蛋白复合体。具体来说，我们建议 致： 1.确定钙粘附素在细胞运动中的作用。 2.描述β-连环蛋白在结构和功能上的差异 和白蛋白(γ-连环蛋白)。 3.检查α-连环蛋白是否有额外的结合蛋白。 为了实现这些目标，我们将利用分子结构，通常 由嵌合分子组成。这些构建物将被转化为 哺乳动物组织培养细胞观察其对细胞的影响 行为和表型。蛋白质之间的相互作用将由以下因素决定 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。 蛋白质之间的相互作用也将通过共转染进行分析。 两种蛋白质在真核细胞中的相互作用结构域 在完整细胞中的定位(免疫荧光)和共标记 细胞提取物的免疫沉淀。

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.