Cleft palate (CP) is one of the most common birth defects. It may not be life threatening but many functions, such as feeding, digestion, speech, middle-ear ventilation, hearing, respiration, and facial and dental development, can be disturbed because of the structures involved. These problems can also cause emotional, psychosocial, and educational difficulties. It imposes a tremendous health burden and often leaves lasting disfigurement. In humans and mice, the secondary palate forms from outgrowths of neural crest-derived mesenchyme covered with a double layer of epithelial cells. The shelves elevate over the tongue and grow toward each other. The medial edge epithelium (MEE) adheres to form the medial epithelial seam (MES). MES cells then undergo epithelial to mesenchymal transition (EMT), cell apoptosis or migrate to the oral and nasal surfaces to form a mesenchymal cell confluence. This fusion process requires transforming growth factor ?3 (TGF?3), and blocking the expression of this protein or its downstream signaling cascade results in CP. Eph receptors tyrosine kinases and their ephrin ligands are responsible for multiple developmental events such as adhesion and migration. Binding of ephrins to Ephs on opposing cells causes tyrosine kinase activation in the Eph- bearing cells (forward signaling), while binding of Ephs can activate intracellular signaling inside ephrinbearing cells (reverse signaling). Activation of ephrin reverse signaling in chicken palates induces fusion, and it requires phosphatidylinositol-3 kinase (PI3K). Blockage of reverse signaling inhibited TGF?3 induced fusion in the chicken and natural fusion in the mouse palate. Thus, ephrin reverse signaling is necessary to induce palate fusion independent of TGF?3. EMT is orchestrated by a complex network of signaling molecules and it is a critical step for palatal fusion. TGF? family is a multifunctional cytokine that oversees and directs all aspects of cell development, differentiation and survival of essentiall cell types and tissues. Also, it is a suppressor of cell growth and proliferation particularly in tumor cells of epithelial and mesenchymal origins. Ephrin signaling promotes elevation of TGF? signaling. These findings lead to the central hypothesis that the TGF? and Eph/ephrin pathways cooperate in EMT in palatal fusion. Thus, the goal of this research project is to use the palate model system to generate cellular responses and changes to study the basic mechanisms that control EMT during palatogenesis. Therefore the aims of this work are as follows: a) Determine if Eph and ephrins play a role in palatal fusion and b) Establish if ephrin reverse signaling is necessary and sufficient to induce EMT in palatal fusion independent of TGF?.
