Identifying the molecular mechanism for instructed determination of phenotype through reciprocal epithelial-mesenchymal interaction remains a significant problem area in craniofacial biology. The hypothesis being tested is that constituents of the extracellular matrix regulate odontogenic cell behavior resulting in morphogenesis and cytodifferentiation during epithelia-mesenchymal interactions in tooth organogenesis. Neural crest derived matrix constituents provide multiple inductive signal to enamel organ epithelia which results in epithelial morphogenesis and tissue specific amelogenin gene expression. The signals are multiple and are restricted in time and position during organogenesis. The signals are received by the epithelium via transmembrane glycoproteins serving as receptors, enzymes and/or ligands. Transduction of these signal results in the transcription and translation of cell specific gene products. The time and position- restricted synthesis and release of paracrine factors by mesenchymal cells of the odontogenic apparatus, the transduction of signals by the epithelium and the subsequent phenotypic responses all operate in tooth organs maintained in chemically defined culture conditions. The development of high resolution in situ hybridization histochemistry and data analysis including three-dimensional reconstruction of developing mouse molar tooth organs clearly provide feasibility for the proposed studies. We will critically test our hypothesis through the acquisition of the following data: i) Identify the time- and position- restricted expression of amelogenin, preproEGF, beta 1-4 galactosyltransferase (GT) and integrin gene products during mouse molar organogenesis in vivo and in vitro; ii) Produce three dimensional reconstructions of in vivo and in vitro molar tooth organogenesis integrating in situ hybridization histochemistry, immunolocalization and position of cell division data sets; iii) Determine when the genome of inner enamel epithelium become stabilized for amelogenin gene expression in the absence of dental papilla ectomesenchyme; iv) Determine which ECM constituent within the biomatrix contains a motif(s) which induces amelogenin gene expression; v) Determine how the ECM constituent induces amelogenin gene expression.
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