Diseases of the retina result in impairments of sight ranging from loss of visual acuity to total blindness as a result of degenerated or otherwise nonfunctional retinal tissue. The retina develops from a set of multipotent progenitor cells, the specification, proliferation, and differentiation of which is dependent on a set of conserved genes. One of these genes, the retinal homeobox gene, Rx, is essential for regulation of retinal cell progenitor specification and proliferation. Even so, the genetic and molecular details of Rx activation and function are poorly understood. The long-term objective of this application is to gain an understanding of the role of Rx in eye development by elucidating the molecular mechanisms of Rx function and the components of the Rx genetic pathway. The following experimental approaches will be undertaken to achieve this objective. (1) To gain a better understanding of the genetic events leading to the activation of Rx, cis-acting elements and trans-acting factors that regulate Rx promoter activity will be identified and characterized. (2) To elucidate the molecular mechanism of Rx function, the involvement of conserved peptide domains encoded by Rx will be investigated in the context of an intact Rx-responsive promoter. (3) To gain a better understanding of Rx function at different phases of eye development, Rx target genes will be identified and characterized. A collection of candidate Rx target genes will be assembled by chromatin immunoprecipitation (CHIP) and a bioinformatics-based approach. The collection will then be refined to identify authentic Rx targets. The results of these experiments will lead to elucidation of upstream and downstream components of Rx genetic and biochemical pathways, which is essential to our understanding of the biological basis of eye development and disease. Additionally, identification of Rx genetic pathway components will expand the pool of candidate genes involved in eye development and human ocular birth defects. Finally, understanding the mechanisms by which Rx is involved in retinal progenitor cell specification and proliferation has profound implications for our abilities to manipulate and exploit retinal stem cells for novel therapeutic approaches involving the replacement of damaged or nonfunctional retinal tissue.
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