Thursday, April 25, 2024
4/25/2024
Project Summary/Abstract Gene regulation, which takes place at both transcriptional and post-transcriptional levels, plays important roles in retinal development and function. So far, studies of gene regulation in the retina have largely focused on the transcriptional level. Although post-transcriptional mechanisms also are critically involved in various biological processes, little is known about how post-transcriptional regulation impacts retinal development and function. In this project, we propose to address this issue by studying two members of the TTP (tristetraprolin) mRNA binding protein family, Zfp36l1 and Zfp36l2 (collectively referred to as Zfp36l1/2). TTP proteins are CCCH zinc finger proteins highly conserved through evolution, and are involved in diverse biological processes. They carry out their functions by binding to the AU-rich elements (AREs) in the 3’ UTR of target mRNAs to promote their decay. We discovered that Zfp36l1/2 were highly expressed in retinal progenitor cells (RPCs) during development and Müller glial cells and photoreceptors in the mature retina. Further, we have created retina- specific knockout mice of the two genes. Our preliminary analysis of the mutant retinas revealed that single knockout retinas appeared largely normal, but the double knockout (DKO) retina had defects in development and degenerated postnatally. Whereas RPCs give rise to all retinal cell types, Müller glial cells are considered quiescent RPCs in the mammalian retina. Thus Zfp36l1/2 likely play shared roles in these two cell types. Our finding that Zfp36l1/2 were also expressed in photoreceptors indicated that the two proteins may also be directly involved in photoreceptor maintenance. Based on these considerations, we hypothesize that regulation of mRNA decay plays essential roles in the retina, and that Zfp36l1/2 are two critical regulators of mRNA decay functioning redundantly in both retinal development and maintenance. To test this hypothesis, we propose to study the function of these two proteins in the retina by investigating how deletion of Zfp36l1 and Zfp36l2 affects both the development and maintenance of the retina, and by dissecting the genes and pathways controlled by Zfp36l1/2 at different developmental stages and in different cell types using a combined approach of mouse genetics, histology, immunofluorescence, electrophysiology, RNA-seq and single cell RNA-seq, CLIP (cross-linking immunoprecipitation)-seq, and bioinformatics. The results from these experiments collectively will allow us to uncover how Zfp36l1/2 are involved in retinal development and maintenance, to identify mRNA targets and relevant pathways regulated by them, and to reveal the shared and unique mechanisms by which these two proteins function at different developmental stages. Therefore, this project affords a unique opportunity to advance our understanding of the roles mRNA decay plays in both normal and disease conditions, and the discoveries we make will add a new dimension to our knowledge of gene regulation in the retina.
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