Development and function of corneal lens-secreting cells - The human cornea is a precisely curved transparent structure composed primarily of extracellular matrix (ECM) that is able to refract light and focus it on the retina. Genetic defects in corneal development contribute to visual disorders such as myopia, hyperopia and astigmatism, as well as more serious diseases like keratoconus and corneal dystrophy. The power of genetic model organisms has been under-used to understand corneal disorders because of a dearth of information about potentially appropriate models such as the Drosophila corneal lens. The corneal lens is entirely made up of apical ECM that is secreted by a small set of non-neuronal retinal cells. These cone and pigment cells develop from the same pool of progenitor cells as photoreceptors, and express the same transcription factor, Glass. The process by which cells in the retina diversify and execute different developmental programs is not well understood. In the first aim of this proposal, Glass-dependent enhancers specific for cone and pigment cells will be combined with a photoreceptor-specific enhancer to determine whether cell type-specific gene expression relies on coactivators or on repressors. These other transcription factors will be identified by extracting their binding motifs from a genome-wide set of cone and pigment cell Glass-bound enhancers obtained from Targeted DamID experiments. Altering the normal timing of cone and pigment cell differentiation, which is controlled by the steroid hormone ecdysone, disrupts corneal lens morphology. Preliminary data suggests that two ecdysone-regulated transcription factors, Blimp-1 and Eip93F, have opposite effects on the progression of differentiation. In the second aim, this hypothesis will be tested by looking for temporal changes in gene expression in the retina in loss of function conditions for either or both genes. In addition, cell type-specific Omni-ATAC will be used to examine how these transcription factors affect chromatin accessibility on cone and pigment cell genes. These experiments will reveal how cell differentiation is precisely timed so that corneal lens components can be deposited in the correct sequence. Finally, a major roadblock to using the corneal lens as a model is the current lack of knowledge about the nature and organization of these components. The third aim will begin to address this by building on recent advances in the transcriptomics of retinal cells and in the biology of other cuticular structures. Components predicted to localize to different layers of the corneal lens will be labeled with endogenous fluorescent tags and used to characterize the organization of the corneal lens in wild-type and mutant conditions. In addition, mutations in genes that encode major corneal lens components will be generated and analyzed to determine their effects on its structure. These studies will reveal which molecular and structural homologies link the Drosophila corneal lens and the human cornea, and will therefore provide an informed basis for using the corneal lens to understand genetic diseases of the cornea and to devise potential treatments for them.
