Cell-type– and developmental stage–specific regulation of gene expression in the retina

项目摘要 在视网膜发育过程中，超过8,000个基因的表达发生了变化， 视网膜祖细胞在进化保守的出生中产生7类细胞类型中的每一类 秩序尽管已经很好地确定，对DNA的共价修饰的变化和 组蛋白和高阶DNA环伴随着基因表达的变化， 这些过程在视网膜发育过程中是如何协调的。在过去的五年里，我们发展了 一个详细的人类和小鼠视网膜基因组的结构和可访问性的地图， 发展具体来说，我们进行了多方面的综合分析，包括分析 对DNA和组蛋白的共价修饰，启动子结构，染色质可及性，成环 相互作用和常染色质/异染色质定位。所有这些已发表和未发表的数据 通过我们的集成视网膜核组与生物医学研究社区免费共享 数据库（iRNDb）（https：//pecan.stjude.cloud/retinalnucleome）。最重要的发现之一 来自iRNDb的是一系列核心调控回路超级增强子的鉴定 （CRC-SE）邻近在视网膜发育中具有重要作用的基因，包括Vsx 2，Crx，Six3， Otx2、Fgf15和Ascl1。Vsx2基因上游的CRC-SE特别令人兴奋，因为它 具有与双极细胞发育一致的活性。我们在小鼠中删除了Vsx2-CRC-SE， 显示双极神经元不存在，但所有其他细胞类型发育正常。重要的是，视网膜 祖细胞增殖是正常的，表明我们已经分离了双极细胞调节， 从视网膜祖细胞中分离出来。在本提案中，我们将阐明结构和 组织Vsx2 CRC-SE，鉴定可能与Vsx2合作的其他转录因子， 调节双极细胞类型特异性表达，并测试双极细胞丧失对 视网膜中的其他细胞。这些研究的结果对于填补一个根本性的空白将是重要的 我们对CRC-SE在视网膜发育中的作用的了解，并将为 在视网膜发生所需的其他基因中的CRC-SE的表征。所有出版和未出版的 通过iRNDb共享数据，以加速对视网膜发育和疾病的发现。

PROJECT SUMMARY During retinal development, more than 8,000 genes change in their expression as multipotent retinal progenitor cells produce each of the 7 classes of cell types in an evolutionarily conserved birth order. Although it has been well established that changes in the covalent modifications to the DNA and histones and higher-order DNA looping accompany changes in gene expression, little is known about how those processes are coordinated during retinal development. Over the past 5 years, we developed a detailed map of the structure and accessibility of the human and mouse retinal genome during development. Specifically, we performed a multifaceted integrated analysis that included profiling of the covalent modifications to the DNA and histones, promoter structure, chromatin accessibility, looping interactions, and euchromatin/heterochromatin localization. All these published and unpublished data are shared freely with the biomedical research community through our integrated retinal nucleome database (iRNDb) (https://pecan.stjude.cloud/retinalnucleome). One of the most significant discoveries to come from the iRNDb was the identification of a series of core regulatory circuit super-enhancers (CRC-SEs) adjacent to genes having important roles in retinal development, including Vsx2, Crx, Six3, Otx2, Fgf15, and Ascl1. The CRC-SE upstream of the Vsx2 gene was particularly exciting because it had activity consistent with bipolar cell development. We deleted the Vsx2-CRC-SE in mice and showed that bipolar neurons are absent yet all other cell types develop normally. Importantly, retinal progenitor cell proliferation was normal, indicating that we had separated the bipolar cell regulatory elements from that of retinal progenitor cells. In this proposal, we will elucidate the structure and organization of the Vsx2 CRC-SE, identify other transcription factors that may cooperate with Vsx2 to regulate bipolar cell type–specific expression and test the consequences of loss of bipolar cells on other cell types in the retina. The results of these studies will be important for filling a fundamental gap in our knowledge about the role of CRC-SEs in retinal development and will set the stage for characterization of CRC-SEs in other genes required for retinogenesis. All published and unpublished data are shared through the iRNDb to accelerate discovery on retinal development and disease.