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

我们的长期目标是了解晶状体发育的机制。在这项应用中，我们更受限的目标是确定不寻常的跨膜蛋白CRIM1如何在发育中的晶状体中介导上皮黏附和形态发生。我们的初步分析表明，CRIM1通过与β-连环素和钙粘附素的相互作用来调节细胞黏附。CRIM1可以与b-连环蛋白形成复合体，进而与钙粘附素相互作用。我们的中心假设是，CRIM1通过与连环蛋白和钙粘附素形成复合体，调节上皮细胞的黏附和形态发生。为了研究CRIM1在晶状体中的功能，我们提出了两个特定的目标：目的1.确定CRIM1、β-连环蛋白和钙粘附素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分析，我们将确定CRIM1是否与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，因为已知这会影响其他β-连环蛋白配体的结合。我们还将确定CRIM1-β-连环蛋白/钙粘连蛋白复合体的化学计量比，因为这可能具有重要的生物学意义。目的2.确定CRIM1-β-catenin相互作用是否调节晶状体中钙粘附素依赖的黏附。使用不能或必须与β-连环蛋白相互作用的CRIM1形式，我们将通过挽救CRIM1条件突变小鼠来确定CRIM1-β-连环蛋白相互作用是否对发育中的晶状体中钙粘附素依赖的黏附重要。发育形态发生的研究本身很有趣，但对我们对疾病的理解也有重要的影响。例如，钙粘附素粘附性的调节对于眼部特有的疾病至关重要，如继发性白内障的形成，在这种疾病中有上皮向间充质的转变，以及在需要血管内皮细胞VE-钙粘附素调节的血管疾病中，糖尿病视网膜病变和年龄相关性黄斑变性。钙粘附素的表达和活性下调也被认为是上皮性肿瘤进展的关键步骤，因为它允许肿瘤细胞离开上皮细胞，成为侵袭性的，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.