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.