BONE GENERATION AND REGENERATION

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The long-term goal of the proposed project is to understand the molecular mechanisms of bone generation and regeneration. During investigations of the involvement of cartilage matrix molecules in alveolar bone repair, the temporal expression of the short form of type IX collagen was demonstrated during the early healing stages of the tooth extraction socket. The alternative selection of two distinct promoters in the (alpha-1(IX) gene can generate long type IX collagen that is transcribed from the upstream promoter contains a large amino-terminal globule, and short type IX collagen that is transcribed from the downstream promoter completely omits the globule. The spatial localization of short type IX collagen in the tooth extraction socket was found mostly in the relatively unorganized matrix and the actively forming bone trabeculae. The extraction socket of alpha-1(IX) "knock-out" transgenic mice showed retarded and disorganized bone repair. Therefore. we hypothesize that short type IX collagen may have a significant role in alveolar bone repair and possibly in any bone formation processes. As a member of the FACIT class of matrix molecules, type Ix collagen decorates the surface of type II collagen fibrils and thus provides the site for tissue specific molecular interactions. We. therefore. hypothesize that heterogeneity in type IX collagen molecules due to the alternative promoter selection may be closely related to differential expressions of cartilage and bone phenotypes. The specific goals of the projects proposed in this application are: 1) to investigate the role of short type IX collagen during bone formation, and 2) to understand the molecular mechanism of the tissue specific selection of the alternative promoters in the alpha-1 (IX) gene. To reach these goals, we propose to complete the analysis of the temporal and spatial expression of type II and IX collagens during alveolar bone repair as well as during embryonic bone formation. We will apply molecular biologic and immuno-identification techniques. The specific role of short type IX collagen during alveolar bone repair will be investigated in transgenic mice with inactivated ("knock-out") alpha-1 (XI) alleles. Furthermore, the alternative promoter regions of the alpha- l(IX) gene will be thoroughly characterized. The regulatory mechanism of the alternative transcription will be addressed by reporter gene assay, mobility shift assay and DNA foot printing assay. The findings from these studies will provide further understanding of bone generation and regeneration, and the essential prerequisites for sound healing of bone wounds.