Regulation of interneuron formation in the developing retina

摘要/项目摘要 视网膜疾病或损伤导致视力受损或失明是人类健康问题，可减少 提高生活质量，产生巨大的人力和经济成本。基因治疗与干细胞研究进展 生物学使视网膜修复成为一个可行的目标。尽管如此，合理的修复策略受到以下因素的限制 视网膜生物学和发展的最新知识。视网膜的复杂细胞组成，比如 哺乳动物中枢神经系统的其他部分，是视网膜功能的重要组成部分 能力，但仍未完全了解。哺乳动物的视网膜包括100多种不同的类型 神经细胞的。视网膜发育过程中这种细胞多样性的产生在一定程度上取决于序列 转录调节因子和其他因素的级联作用。我们发现miR-216B microRNA可以 影响视网膜发育，包括无长突和双极中间神经元的形成。我们确定了一个 这种microRNA的靶基因是叉头家族中的转录抑制因子Foxn3，当被抑制时 在发育中的视网膜中使用RNAi或CRISPR可以增加无长突细胞的形成，当过度表达时 减少无长突细胞的形成。Foxn3在视网膜发育过程中的靶基因尚不清楚。这里 我们建议确定在发育中的视网膜中受Foxn3功能丧失或获得调节的mRNAs。我们 我将分析mRNA表达的变化，以确定Foxn3调节视网膜的分子途径 细胞命运的决定。此外，我们将确定Foxn3蛋白在发育过程中结合的基因组位置。 视网膜来寻找候选的靶基因。我们还建议分析在没有miR的情况下视网膜的发育。 216b或相关的miR-216a microRNA，以确定这些miRNAs是否为无长突细胞所必需 分化或其他功能，并评估两个miR-216a/b基因变体的功能，这两个基因变体可能是 与视网膜疾病有关。这些研究将为发育和细胞的调控提供新的见解 哺乳动物视网膜中的测定。预计他们将提供可能与以下内容相关的信息 视网膜疾病，这可能有助于修复视网膜组织的新策略。

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.