The eye lens is a transparent tissue that refracts and focuses light on the retina to allow clear vision. If the lens loses its transparency, vision is impaired, and the disease is termed ?cataract?. Cataract is the major cause of blindness worldwide, commonly found in aged individuals (about half of Americans aged 80 or older develop cataract), but can also be present at birth or develop in early childhood ? termed pediatric cataract. Genetic anomalies are estimated to account for about 25-50% of pediatric cataract cases, and the etiology of majority of these are unknown. Importantly, while the role of signaling and transcription factors in lens development and cataract have been well defined over the last few decades, that of proteins involved in post-transcriptional gene expression control is grossly understudied ? indeed, only four such proteins have been characterized so far in the lens. Our efforts in characterizing these proteins have demonstrated that post-transcriptional regulatory mechanisms are critical for lens development, and their loss results in early onset cataract and eye defects. Identification of these factors was made possible by our novel bioinformatics approach iSyTE (integrated Systems Tool for Eye gene discovery), which is effective in identifying high-priority target genes linked to lens development and cataract. We have now used iSyTE to identify a new post-transcriptional regulator in the lens, namely, Elavl1 (Embryonic lethal abnormal vision (ELAV) like RNA-binding protein). The function of Elavl1 has not been examined in the lens. Therefore, we developed a new Elavl1-targeted lens-specific conditional knockout (KO) mouse model and find that Elavl1cKO mice exhibit early onset eye defects namely, cataract and microphthalmia. In this proposal, we will test the overarching hypothesis that Elavl1 mediates post-transcriptional gene expression control over key regulators of lens development, disruption of which causes cataract and microphthalmia. Specifically, we will address the following goals.
(Aim 1) Characterize the pathogenesis of lens defects in Elavl1cKO mice and gain insights into the structural and molecular underpinning of these defects by comparative analysis of lens morphology, transcriptome and proteome.
(Aim 2) Test the mechanism of Elavl1- mediated control of post-transcriptional gene expression in the lens. Specifically, we will investigate the molecular mechanism of Elavl1 function in control of the key lens development/differentiation transcription factors Pax6, c-Maf and Prox1 among other targets. Identify direct RNA targets of Elavl1 by RNA-immunoprecipitation followed by RNA-Sequencing.
(Aim 3) Examine how Elavl1 and Celf1 coordinately mediate post-transcriptional control in the lens and integrate and analyze all the molecular, genomic and functional data generated above to derive Elavl1 and Celf1-regulatory networks in the lens. The expected overall impact of these innovative investigative approaches aimed at uncovering the mechanism of Elavl1 function is that it will continue to advance our knowledge of gene expression regulation in the lens at the post-transcriptional level while informing on the nature of RBP-based combinatorial control, in turn leading to identification of new targets linked to cataract.
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, homeostasis and pathogenesis and will continue development of a publicly available web-based interactive tool ? for both clinicians and scientists ? that facilitates prediction and prioritization of genes/pathways to further accelerate cataract gene discovery.
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|Srivastava, Rajneesh; Budak, Gungor; Dash, Soma et al. (2017) Transcriptome analysis of developing lens reveals abundance of novel transcripts and extensive splicing alterations. Sci Rep 7:11572|
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|Wang, Yichen; Terrell, Anne M; Riggio, Brittany A et al. (2017) ?1-Integrin Deletion From the Lens Activates Cellular Stress Responses Leading to Apoptosis and Fibrosis. Invest Ophthalmol Vis Sci 58:3896-3922|
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|Cavalheiro, Gabriel R; Matos-Rodrigues, Gabriel E; Zhao, Yilin et al. (2017) N-myc regulates growth and fiber cell differentiation in lens development. Dev Biol 429:105-117|
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