Cataract, the opacification of the ocular lens, is the leading cause of blindness worldwide. The etiology of most cases of sporadic pediatric cataract is still unknown, and genetic anomalies are estimated to account for 25-50% of these cases. We have developed a novel bioinformatics approach, iSyTE (integrated Systems Tool for Eye gene discovery, http://bioinformatics.udel.edu/Research/iSyTE) to predict genes that are associated with cataract. In the recent past, our finding that deficiency of the RNA granule (RG) component TDRD7 causes juvenile cataracts in human and animal models have indicated the significance of post-transcriptional regulatory mechanisms in the development and maintenance of lens transparency. We have now used iSyTE to identify a new conserved RNA binding protein and RG component Celf1 whose germline or lens-specific deletion in mouse mutants causes early onset cataracts. In this proposal, we will test the overarching hypothesis that Celf1 mediates post-transcriptional control of gene expression to regulate lens development. Specifically, we will address the following goals.
(Aim 1) Characterize the pathogenesis of lens defects in Celf1 mouse mutants and gain insights into the molecular underpinning of lens defects in Celf1 mutants by analysis of the lens transcriptome and proteome.
(Aim 2) Elucidate the direct RNA targets of Celf1 by RNA-immunoprecipitation followed by RNA-Sequencing (RNA- Seq). Further, test the mechanism of Celf1-mediated regulation for high-priority candidate genes that function in the lens, and are selected based on stringent filtering criteria. Specifically, we will investigate the molecular mechanism of Celf1 function in: 1) translational control of the cell cycl kinase inhibitor p27, 2) control of Prox1 expression in lens epithelium, and 3) control of mRNA stability for the fiber cell differentiation factor essential for nuclear degradation, Dnase2b. (Ai 3) Finally, integrate and analyze these data on Celf1 within the context of existing lens data to derive Celf-regulatory networks in the lens. These investigations aimed at elucidating the mechanism of Celf1 function will advance the understanding of post-transcriptional control of gene expression in the lens and lead to identification of new targets associated with cataracts.
Cataracts are the leading cause of blindness worldwide. Therefore, it is essential to understand the molecular mechanisms that contribute to the formation and maintenance of lens transparency. The proposed research will result in the characterization of novel pathways in lens development and pathogenesis and will contribute to a publicly available web-based interactive tool - for both clinicians and scientists - that helps predict and prioritize genes to accelerate cataract gene discovery.
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