PDF(Pubmed)
Abstract：
Desmosomes are large, macromolecular protein assemblies that mechanically couple the intermediate filament cytoskeleton to sites of cadherin-mediated cell adhesion, thereby providing structural integrity to tissues that routinely experience large forces. Proper desmosomal adhesion is necessary for the normal development and maintenance of vertebrate tissues, such as epithelia and cardiac muscle, while dysfunction can lead to severe disease of the heart and skin. Therefore, it is important to understand the relationship between desmosomal adhesion and the architecture of the molecules that form the adhesive interface, the desmosomal cadherins (DCs). However, desmosomes are embedded in two plasma membranes and are linked to the cytoskeletal networks of two cells, imposing extreme difficulty on traditional structural studies of DC architecture, which have yielded conflicting results. Consequently, the relationship between DC architecture and adhesive function remains unclear. To overcome these challenges, we utilized excitation-resolved fluorescence polarization microscopy to quantify the orientational order of the extracellular and intracellular domains of three DC isoforms: desmoglein 2, desmocollin 2, and desmoglein 3. We found that DC ectodomains were significantly more ordered than their cytoplasmic counterparts, indicating a drastic difference in DC architecture between opposing sides of the plasma membrane. This difference was conserved among all DCs tested, suggesting that it may be an important feature of desmosomal architecture. Moreover, our findings suggest that the organization of DC ectodomains is predominantly the result of extracellular adhesive interactions. We employed azimuthal orientation mapping to show that DC ectodomains are arranged with rotational symmetry about the membrane normal. Finally, we performed a series of mathematical simulations to test the feasibility of a recently proposed antiparallel arrangement of DC ectodomains, finding that it is supported by our experimental data. Importantly, the strategies employed here have the potential to elucidate molecular mechanisms for diseases that result from defective desmosome architecture.
摘要：
Desmosomes很大,将中间丝细胞骨架与钙粘蛋白介导的细胞粘附位点机械偶联的大分子蛋白质组装体，从而为经常经受大的力的组织提供结构完整性。适当的桥粒粘附是脊椎动物组织正常发育和维持所必需的。如上皮和心肌，而功能障碍会导致严重的心脏和皮肤疾病。因此,了解桥粒粘附与形成粘附界面的分子结构之间的关系非常重要，桥粒钙粘蛋白（DC）。然而,桥粒嵌入两个质膜，与两个细胞的细胞骨架网络相连，给传统的DC建筑结构研究带来了极大的困难，产生了相互矛盾的结果。因此,DC结构与粘附功能之间的关系尚不清楚.为了克服这些挑战，我们利用激发分辨荧光偏振显微镜来量化三种DC同工型的胞外结构域和胞内结构域的取向顺序:桥粒糖蛋白2,桥粒糖蛋白2和桥粒糖蛋白3.我们发现DC胞外域比它们的细胞质对应物明显更有序，表明质膜相对侧之间的DC结构存在巨大差异。这种差异在所有测试的DC中是保守的，这表明它可能是桥粒结构的重要特征。此外，我们的发现提示DC胞外域的组织主要是细胞外粘附相互作用的结果.我们采用方位角方向映射来显示DC胞外域围绕膜法线旋转对称排列。最后,我们进行了一系列的数学模拟，以测试最近提出的反并联布置的DC域的可行性，发现它得到了我们实验数据的支持。重要的是,这里采用的策略有可能阐明由桥粒结构缺陷引起的疾病的分子机制。
