Cell-Cell Signaling During Mammalian Early Eye Formation

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DESCRIPTION (provided by applicant): This proposal investigates the underlying causes of human ocular diseases using mouse models, focusing on the Notch signaling pathway, which is broadly required during development. Notch signaling regulates proliferation, cell shape changes, differentiation, and apoptosis and stem cell maintenance in specific cellular contexts. Dominant human mutations in the ligand Jagged1 or Notch2 receptor cause Alagille syndrome, in which some patient's exhibit eye deformities. In the last funding period we discovered a Jag1- Notch1/2-Rbpj-Hes1 core signal is required for prenatal mouse lens formation. Mice lacking each of these genes exhibited progressive postnatal lens aphakia, loss of the pupillary opening and microphthalmia. Using germline or conditional mutant alleles, and deletion of two or more genes in complex mutants, we propose to investigate how Presenilin (Psen) proteases regulate Notch receptor activation to release a c-terminal polypeptide (NICD) that complexes in the nucleus to regulate downstream genes. Additional Notch-independent roles for Psen genes will also be explored. Moreover, the Notch ligands Dll1, Jag1 and Jag2 are posttranslationally regulated by ubiquitination. The addition of one or more ubiquitin molecules targets the ligands for degradation, or endocytic recycling within signal sending cells. We propose to explore the role of two families of E3 ubiquitin ligases, Neurl and Mib genes in the developing lens, for which nothing is currently known. Finally we will determine the independent, and combined, roles of the Hes1 and Hes5 downstream effector genes, during Notch regulation of growth versus differentiation. The studies described in this application will extend our understanding of how Notch promotes lens progenitor cell proliferation, promotes fiber cell differentiation and prevents inappropriate fiber cell mitoses or excess apoptosis. All experiments will employ complex conditional (cre-lox) mouse strains, including double and triple mutants, histology, immunohistochemistry, confocal microscopy, in situ hybridization, mouse embryology, Western blotting, qPCR and PCR genotyping. We hope to contribute new information to the processes of growth, morphogenesis and differentiation, which are fundamental to all cells and tissues.