Studies on the Structure of Basement Membranes

COL4A3、COL4A4和COL4A5基因中的数百种变体导致广泛的肾小球疾病 影响肾小球基底膜（GBM）的功能。这些基因编码的组装 胶原IV α345支架，GBM的主要成分，Goodpasture（GP）中的自身抗原 自身免疫性疾病，以及Alport综合征和其他遗传性肾小球病中突变的蛋白质。GP 疾病已经并将继续作为解开细胞分子结构之谜的先锋， α345支架和获得性和遗传性肾小球病的致病机制我们 总体假设是：胶原蛋白IV α345支架束缚大分子形成超分子 支架的复合物和扰动引起肾小球病。四个具体目标解决关键问题 根据我们以前和最近的发现定义的未回答的问题。目标1：α 345NC 1 六聚体确定α 345NC 1六聚体的原子结构和GP表位的作用机制 阵GP自身抗原的结构是未知的。我们假设， 非免疫原性的EST345NC1六聚体、EA和EB区的四级结构的扰动 经历构象变化，形成致病性GP新表位。目的2：α3苏黎世突变。到 确定α3苏黎世突变对GP表位形成的影响。我们在α 3NC 1基因中发现了一个突变， 与第一例家族性GP疾病相关的结构域，为触发GP疾病提供了遗传证据。 机制我们假设突变引起了Ea和EB区域的结构扰动， α 3NC 1参与GP表位的呈递。目标3。氯环。确定的作用 氯化物在胶原IV α345支架组装和GP表位形成中的作用。而结构， α121支架的组装和功能已经成功地研究了40多年，我们的知识 关于α345支架的信息仍然不清楚。我们证明了氯离子浓度是GP的关键因素 抗体结合我们假设，在这种情况下，HPL345NC1六聚体的组装、其稳定性和GP反应性可能是一个重要的因素。 依赖于氯离子。目的4：α121超分子配合物。为了表征超分子 基底膜内的α121胶原IV复合物。我们发现了一个加兰结构 α121蛋白多糖包被支架。这种上层结构是基底膜的潜在核心特征。 我们假设IV型胶原α121支架栓系大分子，形成不同的超分子 使基膜组装成为可能的复合物。目标的实现将产生新的见解 GP疾病的病因学和IV型胶原支架的结构和组装，导致一个框架 用于开发GBM疾病的新治疗策略。

Hundreds of variants in the COL4A3, COL4A4 and COL4A5 genes cause a broad range of glomerulopathies affecting the function of the glomerular basement membrane (GBM). These genes encode the assembly of collagen IV α345 scaffolds, the major constituent of the GBM, the autoantigen in Goodpasture’s (GP) autoimmune disease, and the protein mutated in Alport syndrome and other genetic glomerulopathies. GP disease has and continues to serve as the vanguard for unlocking mysteries of the molecular structure of the α345 scaffold and pathogenic mechanisms underlying both acquired and genetic glomerulopathies. Our overarching hypothesis is: Collagen IV α345 scaffold tethers macromolecules forming supramolecular complexes and perturbation of scaffold causes glomerulopathies. Four specific aims address key unanswered questions that are defined based on our previous and recent discoveries. Aim 1: α345NC1 Hexamer. To determine the atomic structure of the α345NC1 hexamer and mechanism of GP epitopes formation. The structure of the 345NC1 hexamer, GP autoantigen, is unknown. We hypothesize that upon perturbation of quaternary structure of the non-immunogenic 345NC1 hexamer, EA and EB regions undergo conformational changes forming pathogenic GP neoepitopes. Aim 2: α3 Zurich Mutation. To determine the impact of α3 Zurich mutation on GP epitopes formation. We found a mutation in α3NC1 domain associated with the first case of familial GP disease, providing genetic evidence for a triggering mechanism. We hypothesize that the mutation causes structural perturbation of the EA and EB regions of α3NC1, which can contribute to GP epitopes presentation. Aim 3. Chloride ring. To determine role of chloride in assembly of the collagen IV α345 scaffold and formation of GP epitopes. Whereas structure, assembly and functions of the α121 scaffold has been successfully studied for over 40 years, our knowledge about the α345 scaffold remains obscure. We demonstrated that chloride concentration is a critical factor in GP antibody binding. We hypothesize that assembly of the 345NC1 hexamer, its stability and GP-reactivity is dependent on chloride ions. Aim 4: α121 Supramolecular complexes. To characterize the supramolecular complexes of α121 collagen IV within a basement membrane. We discovered a garland architecture of the α121 scaffold coated with proteoglycans. This suprastructure is a potential core feature of basement membrane. We hypothesize that collagen IV α121 scaffold tethers macromolecules forming distinct supramolecular complexes which enable basement membrane assembly. The achievement of the aims will yield new insights to the etiology of GP disease and the structure and assembly of collagen IV scaffolds, leading to a framework for development of novel therapeutic strategies for GBM diseases.