Type XV is a nonfibrillar collagen discovered from the sequence o cDNA clones. The alpha chain consists of a 577 residue highly interrupted triple-helical region flanked by 555 and 256 residue NH2- and COOH-terminal domains. RNA hybridization studies revealed that type XV transcripts are expressed in many tissues and cell types. A polyclonal antibody recognizing th COOH-terminus was developed and used in light microscopy immunohistochemical examination of ten human tissues. The results showed a consistent and almost exclusive presence of the protein in basement membrane zones, i.e., some epithelial, and all muscle, perineural and endothelial except for the glomerular capillaries. The unusual localization suggested a possible role for this collagen as a prominent and nearly ubiquitous link protein between the basement membrane and underlying connective tissue stroma, comparable to the function of type VII collagen in the few tissues lined by stratified squamous epithelium. Western Blot analysis of human tissue and cell culture extracts provided strong evidence that the type XV trimers incorporated into tissues ar a processed form of a higher molecular weight precursor. This proposal is designed to determine in depth the structure, function and expression of type XV collagen. Native type XV collagen will be purified from human placenta tissue, and its macromolecular structure, including the existence of end-terminal interactions, will be examined by rotary shadowing electron microscopy. NH2-terminal protein sequencing will determine the molecular composition of type XV, and the location of the putative cleavage site expecte to occur at the end of the noncollagenous NH2-terminus. The presumed conversio of the alpha1 (XV) chain from a 210-kDa monomer to the 116-kDa processed form will also be investigated in vitro by biosynthetic time course and pulse-chase studies using cultured human cells and domain-specific antibodies. The ultrastructural organization of type XV collagen in basement membrane zones of human skin, kidney and colon will be explored by immunogold electron microscopy. Incorporated into these studies is the association of type XV with another newly discovered and nearly ubiquitous basement membrane zone matrix molecule, type XIX collagen. Conclusive results from the experiments proposed here will be critical to defining whether a previously unrecognized and prevalent complex exists at the interface of the lamina densa and subjacent layers. The organization and structure of type XV collagen and the molecules with whom it interacts will elucidate their involvement in inherited diseases of connective tissue and other pathologic processes.
