Basement membranes are sheets of extracellular matrix with the potential to influence tissue and cell behavior. They can serve as an adhesion scaffolds, regulate permeability, and influence migration, development and differentiation. Basement membrane composition includes collagens, glycoproteins and proteoglycans. Bamacan is a chondroitin sulfate proteoglycan that may regulate basement membrane stability since its expression is low in developing tissue and disrupted in skin and other organs in diseases where matrix integrity is compromised, e.g. dystrophic epidermolysis bullosa. Bamacan core protein has an unusual five domain head-rod-tail structure, not seen in any other matrix component, consistent with a distinct, but as yet unknown, role in matrix biology. The long term objective of this research is to understand its function in normal and diseased tissues. Molecular, immunochemical and cell culture studies are proposed that target: 1) the structure and organization of bamacan core protein. Protein sequence predicts that the rod domains form coiled-coil alpha helices, with stabilization through dimer formation. A partner protein for bamacan will be identified and characterized. Intermolecular interactions between head and tail domains in homologous or heterologous assemblies will be elucidated. Further proposed experiments examine the sites and role of bamacan's chondroitin sulfate chain substitution. 2) the hypothesis that bamacan influences basement membrane stability. Cell culture systems will be used to examine whether the presence of bamacan affects the rate of matrix turnover. Three different methodologies, incorporating regulated bamacan expression and its glycanation are proposed. 3) bamacan core protein and carbohydrate expression in patients with epidermolysis bullosa and the effects of bamacan on key features of cell behavior including adhesion, cytoskeletal organization and migration. 4) the molecular properties of human bamacn, and the regulation of expresson by keratinocytes. The epidermal compartment is the source of dermal-epidermal junction bamacan in vivo, but this is not matched in vitro, perhaps as seen in disease. Roles for regulation by matrix and growth factors will be assessed. Genomic cloning of mouse bamacan is proposed, with emphasis on the 5' regions, directed towards understanding gene regulation as well as future targeted deletion.
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