A High-Resolution Enhancer Atlas of the Developing Forebrain

描述（申请人提供）：哺乳动物前脑的发育严重依赖于基因表达的动态但精确的空间和时间控制。虽然正在进行的公共努力正在积极地绘制基因组尺度上的基因表达模式，但驱动这些精美模式的转录增强子序列仍未得到明确定义。比较基因组学方法越来越能够相对自信地预测假定的远距离作用增强子的位置，但由于缺乏高质量的实验数据集，对其确切组织特异性的更深入了解尚未出现。在初步研究中，我们将比较基因组学与高通量小鼠转基因报告试验相结合，确定了100多个远程作用增强子序列，这些序列可重复地驱动发育中的前脑的体内表达，并且在许多情况下位于已知的大脑发育所需的基因附近。在这里，我们建议利用这种独特的增强子集合来产生第一代高分辨率的发育前脑顺式调控图谱。这一目标将通过详细的组织学和神经解剖学的分析，所有前脑增强确定到目前为止。然后，我们将收集驱动前脑内相同模式的元件，并计算定义它们的共同转录因子结合位点和其他序列特征，并使用该前脑顺式调控代码生成全基因组增强子目录。这些预测的有效性将通过在转基因小鼠中测试200种这些元素来确定。来自初始图谱的高分辨率数据和注释以及通过我们的预测产生的增强子将通过现有的门户网站提供给社区，使研究人员能够访问这些数据：a)了解前脑基因的调控，b)确定人类遗传疾病中调控突变筛选的候选区域，c)检索用于各种下游实验应用的组织特异性试剂。D)下载数据集用于计算或实验监管研究。了解基因如何控制人类大脑发育的努力主要集中在基因在发育过程中的活跃时间和位置，但在很大程度上忽略了控制这些活动的基因开关（“增强子”）的作用。在这里，我们建议绘制大脑中几百个增强子的精确活动，以帮助预测整个人类基因组中额外的前脑增强子，从而为科学界提供一个“大脑增强子图谱”。这些研究可能会对定义这些开关在正常大脑功能中的作用以及它们在人类大脑疾病中如何出错产生影响。

DESCRIPTION (provided by applicant): Development of the mammalian forebrain critically depends on the dynamic but precise spatial and temporal control of gene expression. While ongoing public efforts are actively mapping gene expression patterns on a genomic scale, the transcriptional enhancer sequences that drive these exquisite patterns remain poorly defined. Comparative genomic methods increasingly enable relatively confident predictions of the location of putative distant-acting enhancers, but a deeper understanding of their exact tissue-specificities has yet to emerge due to the lack of high-quality collections of experimental datasets. In preliminary studies, we have coupled comparative genomics to a high-throughput mouse transgenic reporter assay and identified more than 100 distant-acting enhancer sequences that reproducibly drive in vivo expression in the developing forebrain and are in many cases located near genes known to be required for brain development. Here we propose to exploit this unique enhancer collection to produce a first-generation high-resolution cis-regulatory atlas of the developing forebrain. This goal will be achieved through detailed histological and neuroanatomical analysis of all forebrain enhancers identified so far. We will then bin elements that drive identical patterns within the forebrain and computationally define their common transcription factor binding sites and other sequence features and use this forebrain cis-regulatory code to generate genome-wide enhancer catalogues. The validity of such predictions will be determined through the testing of 200 of these elements in transgenic mice. High-resolution data and annotations from the initial atlas as well as enhancers generated through our predictions will be made available to the community through an existing web portal, allowing researchers to access this data to a) understand the regulation of forebrain genes, b) identify candidate regions for regulatory mutation screens in human genetic disorders, c) retrieve tissue-specific reagents for a variety of downstream experimental applications, d) download data sets for computational or experimental regulatory studies. PUBLIC HEALTH RELEVANCE Efforts to understand how genes control the development of the human brain have focused on when and where genes are active during development, but have largely ignored the role of genetic switches ("enhancers") that control these very activities. Here we propose to map the precise activity of several hundred enhancers in the brain to aid in predicting additional forebrain enhancer in the entire human genome and thereby provide the scientific community with a "brain enhancer atlas". These studies are likely to have implications in defining the role of these switches for normal brain function and how they go awry in human brain diseases.