Extracellular matrix proteins are known to serve not only as structural elements in tissues but also to influence cell shape, movement, biosynthetic phenotype, and proliferation. In the animal, these functions contribute to embryonic development, morphogenesis, and tissue differentiation. Thrombospondin (TSP) belongs to a class of extracellular proteins that is thought to modulate cell-matrix interactions, and may play a role in regulating cell attachment, proliferation, and migration, and in angiogenesis. This laboratory has demonstrated that there are at least three expressed genes coding for related TSPs. The existence of three protein chains that are likely to demonstrate both similarities and crucial differences in structure and function obliges a reevaluation of previous work relating to the location and function of TSP, and greatly increases the potential complexity of TSP-cell interactions and the range of functions which this group of proteins could perform. Our objectives in this proposal are to isolate and characterize the three mouse TSP genes, to compare the pattern of mRNA and protein expression in mouse cells and tissues, and to assess the response of these genes to growth factors and to cytokines. Since mutations in TSPs are likely to exert dominant negative effects, minigenes that exclude several exons will be constructed and introduced into mice. Transgenic mice carrying such genes could demonstrate a phenotype that reflects the function of the mutated TSP. Finally, targeted disruption of the TSP genes will be performed by homologous recombination in somatic cells to evaluate the requirement of the TSPs for cell attachment, migration, and proliferation. It is expected that these experiments will clarify the function of the individual TSPs and address the possibility of the existence of heterotrimeric TSPs. It will be instructive to examine the regulation of the three genes and to relate these findings to the pattern of expression and function of the three proteins. This information should increase our understanding of the role of the TSPs in the response of tissues to injury and, in particular, to the reaction of blood vessel wall to endothelial damage.
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