CRIM1-b-Catenin-Cadherin Interactions in Eye Development and Disease

我们的长期目标是了解透镜开发的机制。在本申请中，我们更严格的目标是确定不寻常的跨膜蛋白CRIM 1如何介导上皮细胞粘附和发育中的透镜的形态发生。我们的初步分析表明，CRIM 1通过与β-连环蛋白和钙粘蛋白的相互作用来调节细胞粘附。CRIM 1可以与b-连环蛋白形成复合物，该复合物又与钙粘蛋白相互作用。我们的中心假设是通过与连环蛋白和钙粘蛋白形成复合物，CRIM 1调节上皮细胞粘附和形态发生。为了研究CRIM 1在透镜中的功能，我们提出了两个具体目标：目标1。探讨CRIM 1、β-catenin和cadherins之间的相互作用机制。通过基于细胞或基于重组蛋白的相互作用测定，我们将确定CRIM 1是否以任何已知的配体结合机制结合β-连环蛋白。这将包括对磷酸化作用的评估，因为已知磷酸化影响其他β-连环蛋白配体的结合。我们还将确定CRIM 1-β-连环蛋白/钙粘蛋白复合物的化学计量，因为这可能具有重要的生物学意义。目标二。确定CRIM 1-β-连环蛋白相互作用是否调节透镜中钙粘蛋白依赖性粘附。使用不能或强制性与β-连环蛋白相互作用的CRIM 1形式，我们将使用CRIM 1条件突变小鼠的拯救来确定CRIM 1-β-连环蛋白相互作用对于发育中的透镜中的钙粘蛋白依赖性粘附是否重要。发育形态发生的研究本身就很有趣，但对我们理解疾病也有重要意义。例如，钙粘蛋白β的调节对于眼睛特异性疾病是至关重要的，例如继发性白内障形成，其中存在上皮向间充质转化，以及在血管疾病糖尿病视网膜病变和年龄相关性黄斑变性中，需要调节血管内皮细胞中的VE-钙粘蛋白。钙粘蛋白表达和活性的下调也是上皮肿瘤进展的关键步骤，因为它允许肿瘤细胞离开上皮，变得具有侵袭性并最终形成转移。

Our long-term goal is an understanding of the mechanisms of lens development. In this application, our more restricted goal is to determine how the unusual transmembrane protein CRIM1 mediates epithelial adhesion and morphogenesis in the developing lens. Our preliminary analysis has shown that CRIM1 regulates cell adhesion through interactions with beta-catenin and cadherins. CRIM1 can form a complex with b-catenin which in turn interacts with cadherins. Our central hypothesis is that through complex formation with catenins and cadherins, CRIM1 regulates epithelial cell adhesion and morphogenesis. To investigate the function of CRIM1 in the lens, we propose two specific aims: Aim 1. To determine the mechanism of interaction between CRIM1, beta-catenin and cadherins. With interaction assays that are cell-based or recombinant protein-based we will determine whether CRIM1 binds beta-catenin with any of the known ligand binding mechanisms. This will include an assessment of the role of phosphorylation as this is known to influence binding of other beta-catenin ligands. We will also determine the stoichiometry of CRIM1-beta-catenin/cadherin complexes as this may have important biological implications. Aim 2. To determine whether the CRIM1-beta-catenin interaction regulates cadherin-dependent adhesion in the lens. Using forms of CRIM1 that either cannot or obligatorily interact with beta-catenin we will determine, using rescue of CRIM1 conditionally mutant mice, whether the CRIM1-beta-catenin interaction is important for cadherin dependent adhesion in the developing lens. The study of developmental morphogenesis is interesting in its own right but also has important consequences for our understanding of disease. For example, regulation of cadherin adhesiveness is critical for eye-specific diseases such as secondary cataract formation where there is an epithelial to mesenchymal transition and in the vascular diseases diabetic retinopathy and age-related macular degeneration that require the modulation of VE-cadherin in vascular endothelial cells. The down-regulation of cadherin expression and activity is also know to be a critical step in epithelial tumor progression as it allows tumor cells to leave the epithelium, become invasive and ultimately form metastases.