The latent TGF-beta binding protein-1 (LTBP-1) is a 120-170 kD molecule with strong homology to fibrillins-1 and -2; the proteins defective in Marfan syndrome and Congenital Contractual Arachnodactyly. LTBP-1 has 18 EGF-like repeats, most of which are Ca++-binding, and 4 repeats of a cysteine-rich domain. Fibrillins-1 and -2 contain over 40 EGF-like repeats and seven cysteine-rich domains. Thus far, 2 cysteine-rich domains have only been described in the LTBPs and fibrillins. Consistent with the widespread in vivo distribution of LTBP-1, in vitro experiments indicate that LTBP-1 is required for TGF-beta release from its latent complex, is necessary for bone nodule differentiation, and is critical for epithelial-mesenchymal transformation during endocardial cushion formation. Thus, LTBP-1 may carry out duel functions. It may be important in matrix organization, and/or it may target latent TGF- beta to sites where subsequent release regulates normal tissue maturation. However, the mechanisms and consequences of LTBP-1 activity in matrix organization and/or in TGF-beta function in vivo have not been elaborated and the differentiation of LTBP-1 activity as either a matrix structural component or as a latent TGF-beta targeting molecule has not been described. We will analyze these possibilities using a genetic approach whereby mice will be developed that express either 1) TGF-beta1 modified so that the cysteine required for bonding to LTBP-1 is mutated to serine, thereby blocking TGF-beta1 interaction with LTBP-1, or 2) LTBP-1 either missing specific sequences necessary for fibrillogenesis or with a null mutation. The phenotypes of animals expressing these mutated proteins will be analyzed, and cells and tissues derived from these mice will be characterized by histologic and functional assays to analyze fibrillogenesis, TGF-beta activation, bone nodule formation, and endocardial-mesenchymal transformation. Studies with mice expressing the TGF-beta mutation should clarify the relationship between LTBP-1 binding and TGF-beta activation and indicate the importance of matrix bound latent TGF-beta. Studies with mutated LTBP-1 should elucidate the role of LTBP-1 in the organization of connective tissue and may reveal a phenotype to suggest a role for LTBP-1 in human disease. The information gained from these studies will reveal the function of LTBP-1 and improve our understanding of tissue organization and growth factor action in both normal and pathological states including cancer, cardiovascular malformation, and bone formation.
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