Creation of gene targeted mutations and the introduction of dominant mutations in transgenic mice are useful for understanding the function of proteins in development and for screening animal models of various diseases for potential therapies. The purpose of this project is to create animal models in mice for studying the molecular basis of genetic and acquire diseases associated with connective tissues. These models will also be useful in elucidating the role of these proteins in development. Genes for the basement membrane and cartilage components have been cloned and the exon-intron structure of these gene have been characterized. Mouse laminin chains and type IV collagen chains have been cloned and sequenced. Several cartilage proteins including aggrecan, link protein and type II collagen have also been cloned and sequenced. The promoters of these genes have been identified and their activity examined. We have used these genes to prepare a gene targeting vector to create gene knockout mice. We have been identifying an enhancer of the genes necessary for tissue- specific expression in transgenic mice. These tissue- specific enhancers and promoters will be used to express foreign genes in specific tissues in a developmental stage-specific manner. These mice would be very useful as an animal model for diseases associated with cartilage and basement membrane. We have characterized naturally occurring mutant mice with connective tissue abnormalities. For example, we have identified a deletion in the aggrecan gene of cmd (cartilage matrix deficiency) mice and a mutation in the laminin alpha2 chain gene of dy (dystrophia muscularis) mice. We have attempted to rescue these mice by introducing the normal genes.