The long-term goal of the proposed study is to elucidate the molecular mechanisms by which signal transduction pathways regulate eye development and ocular surface morphogenesis. Embryonic eyelid closure is essential for normal vertebrate ocular surface morphogenesis; its deficiency in mouse results in eyes open at birth (EOB) and various eye pathologies. Many human congenital diseases, such as conjunctiva-corneal dystrophy, ptosis and microophthalmia, exhibit similarities to pathologies found in mice with EOB and are possibly linked to failure in eyelid development. Understanding the molecular events in eyelid closure may uncover the causes of congenital eye diseases, help develop diagnostic tools and identify targets for possible pharmaceutical intervention. Ablation in mice of the Mekkl gene, encoding a cytoplasmic protein kinase, causes EOB phenotype. MEKK1 ablation results in defects in F-actin formation in developing eyelid epithelium and impairments in keratinocyte cell migration. MEKK1-mediated Jun N-terminal kinase (JNK) cascade leads to phosphorylation of the transcription factor cJun and a concomitant change in gene expression pattern that promotes cell movement and proliferation, but prevents epithelium terminal differentiation. These findings lead to the hypothesis that induction of the MEKK1 pathway in the developing eyelid leads to phosphorylation and activation of members of the AP-1 transcription complex, responsible for changes in gene expression, epithelial cell migration and embryonic eyelid closure. The studies proposed in this application will utilize developing eyelid tissues and cultured keratinoctyes from wild type and Mekk1-/-mice available in our laboratory to investigate the molecular pathways involved in the induction of MEKK1 activity and the regulation of AP-1 function. I propose (1) to elucidate the molecular processes of MEKK1 signaling that lead to keratinocyte migration and mediate eyelid closure; (2) to identify the MEKK1-dependent phosphorylation of AP-1 critical for keratinocyte migration; and (3) to define the role of MEKK1 in the regulation of AP-1 transcription functions pertinent to changes in gene expression during eyelid closure. Results from this work will extend our understanding of the molecular regulation of eyelid development and advance our knowledge on the signaling networks and molecular events critical for epithelial cell migration and eyelid closure.
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