Regulation of interneuron formation in the developing retina

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Abstract/Project Summary Retinal disease or injury leading to impaired vision or blindness are human health problems that reduce quality of life, generating significant human and economic costs. Advances in gene therapy and stem cell biology have made retinal repair a feasible goal. Nonetheless, rational repair strategies are constrained by current knowledge of retinal biology and development. The complex cellular composition of the retina, like other parts of the mammalian central nervous system, is an essential component of the retina’s functional capabilities, yet remains incompletely understood. The mammalian retina includes more than 100 distinct types of neurons. The generation of this cellular diversity during retinal development depends in part on sequential cascades of transcriptional regulators as well as other factors. We found the miR-216b microRNA can influence retinal development including the formation of amacrine and bipolar interneurons. We identified a target gene for this microRNA, a transcriptional repressor in the forkhead family, Foxn3, that when inhibited using RNAi or CRISPR in the developing retina increases amacrine cell formation, and when overexpressed reduces amacrine cell formation. The target genes of Foxn3 during retinal development are not known. Here we propose to identify mRNAs regulated by either loss or gain of Foxn3 function in the developing retina. We will analyze changes in mRNA expression to identify the molecular pathways by which Foxn3 modulates retinal cell fate decisions. In addition, we will identify genomic sites at which the Foxn3 protein binds in the developing retina to find candidate target genes. We also propose to analyze retinal development in the absence of miR- 216b or the related miR-216a microRNA, to determine if these miRNAs are required for amacrine cell differentiation or other functions, and to assess the function of two miR-216a/b genetic variants that may be linked to retinal disease. These studies will provide new insights into the regulation of development and cell determination in the mammalian retina. They are expected to provide information that may be relevant to retinal disease and that may contribute to new strategies to repair retinal tissue.