Genetic Studies of Cortex Structure and Development

皮层结构和发育的遗传学研究

• 批准号: 9214258
• 负责人: JOHN L. R. RUBENSTEIN
• 金额: $ 64.37万
• 依托单位: UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-30 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9214258
• 关键词: Autistic Disorder; Automobile Driving; Binding; Binding Proteins; Biological Assay; CRISPR/Cas technology; Cell Separation; Cell physiology; Cells; Cerebral Palsy; Cerebral cortex; ChIP-seq; Code; DNA Sequence; Development; Embryo; Enhancers; Epilepsy; Fibroblast Growth Factor; Foundations; Gene Expression Regulation; Gene Targeting; Generations; Genes; Genetic; Genetic Enhancer Element; Genetic Transcription; Genetic study; Health; Histones; Human; Human Genetics; Mental Retardation; Mental deficiency; Mitotic; Molecular; Molecular Genetics; Mutation; Nature; Neurons; Pathway interactions; Pattern; Phenotype; Regulator Genes; Regulatory Element; Research; Role; Schizophrenia; Signal Transduction; Structure; Testing; Transcriptional Regulation; Transgenic Mice; Translating; Ventricular; Work; chromatin immunoprecipitation; developmental disease; emx2 protein; epigenomics; genetic information; genetic variant; in vivo; insight; loss of function; loss of function mutation; mutant; nerve stem cell; neuropsychiatric disorder; progenitor; promoter; research study; response; subventricular zone; transcription factor; transcriptome sequencing

PROJECT SUMMARY Dysregulation of the cerebral cortex is central to human developmental disorders such as epilepsy, mental deficiency, autism and schizophrenia. During development, cortical progenitors generate the projection neurons of the different cortical subdivisions. Understanding the genetic circuitry controlling the development and function of these neurons provides an essential foundation for interpreting human allele variants that are enriched in people who have neuropsychiatric disorders. To elucidate this genetic circuitry, we must define the transcription factors (TF), and regulatory elements and of the coding regions that they control. The proposed research, which concentrates on cortical regionalization, involves the systematic identification of TFs, and the regulatory elements and genes downstream of TFs. Currently, the regional-specification functions of a few TFs in embryonic cortical progenitors are known, and little is known about their direct transcription targets, the nature of the regulatory elements that these TFs control, and the transcriptional circuitry that integrates development and function of these cells. Here we propose to make inroads into each of these components of the TF hierarchy regulating cortical development. Furthermore, we aim to elucidate transcriptional mechanisms through which patterning of cortical progenitors is transmitted to, and maintained in, cortical neurons. We hypothesize that enhancers active in the ventricular zone, subventricular zone and the cortical plate are differentially bound by TFs that drive expression of region/layer-specific genes in post-mitotic cortical neurons. The enhancers serve as protein-binding modules that translate rostrocaudal gradients of TFs in cortical progenitors into region-specific expression in cortical neurons. Herein we focus on the transcriptional mechanisms controlling the generation of different regions of the cerebral cortex (cortical regionalization). The Five Specific Aims extend upon our earlier work on regionalization of cortical progenitors by FGF-signaling, TFs and enhancer elements. Here we investigate transcriptional regulation of cortical patterning by defining the TFs, and other genes, that are regulated by COUPTF1, EMX2, and PAX6 (Aim 1). We then use chromatin immunoprecipitation-DNA sequencing (ChIP-Seq) to define the regulatory element (RE) and gene targets of COUPTF1, EMX2, and PAX6 (Aim 2). Next, we use fluorescent activated cell sorting (FACS) to purify cells from the VZ, SVZ, CP and layers 5&6 to elucidate the epigenomic states of REs (and genes) using Histone ChIP-Seq. This will help us understand the molecular mechanisms that transmit regional patterning information from cortical progenitors to neurons (Aim 3). Finally, we define the function of REs related to cortical patterning using transgenic mice to assess RE activity (Aims 4) and REs deletions to define their role in gene regulation (Aim 5). Once integrated with human genetic information, this will enable us to gain powerful insights into how abnormalities in specific gene networks cause human neuropsychiatric disorders.

项目概要 大脑皮层失调是人类发育障碍的核心，例如癫痫、精神障碍等。 缺乏症、自闭症和精神分裂症。在发育过程中，皮质祖细胞产生投射 不同皮质分区的神经元。了解控制发育的遗传电路 这些神经元的功能和功能为解释人类等位基因变异提供了重要基础 患有神经精神疾病的人中含量丰富。为了阐明这种遗传电路，我们必须定义 转录因子（TF）、调控元件及其控制的编码区。拟议的 研究集中于皮质区域化，涉及 TF 的系统识别，以及 转录因子下游的调控元件和基因。目前，一些 TF 的区域规范功能 在胚胎皮质祖细胞中，人们对它们的直接转录目标知之甚少， 这些 TF 控制的调控元件的性质以及整合的转录电路 这些细胞的发育和功能。在这里，我们建议深入研究这些组成部分 调节皮质发育的 TF 层次结构。此外，我们的目标是阐明转录机制 通过它，皮质祖细胞的模式被传递到皮质神经元并维持在其中。我们 假设活跃于心室区、室下区和皮质板的增强子是 被驱动有丝分裂后皮层神经元中区域/层特异性基因表达的转录因子差异结合。 增强子作为蛋白质结合模块，翻译皮质中 TF 的轴尾梯度 祖细胞进入皮质神经元的区域特异性表达。这里我们重点关注转录 控制大脑皮层不同区域生成的机制（皮质区域化）。这 五个具体目标延续了我们早期通过 FGF 信号传导进行皮质祖细胞区域化的工作， TF 和增强子元件。在这里，我们通过定义 受 COUPTF1、EMX2 和 PAX6 调控的 TF 和其他基因（目标 1）。然后我们使用染色质 免疫沉淀-DNA 测序 (ChIP-Seq) 来定义调节元件 (RE) 和基因靶标 COUPTF1、EMX2 和 PAX6（目标 2）。接下来，我们使用荧光激活细胞分选术（FACS）来纯化细胞 使用组蛋白从 VZ、SVZ、CP 和第 5 层和第 6 层阐明 RE（和基因）的表观基因组状态 ChIP 测序。这将有助于我们理解传递区域图案信息的分子机制 从皮质祖细胞到神经元（目标 3）。最后，我们定义了与皮质模式相关的 RE 的功能 使用转基因小鼠评估 RE 活性（目标 4）和 RE 删除以确定其在基因调控中的作用 （目标 5）。一旦与人类遗传信息整合，这将使我们能够深入了解如何 特定基因网络的异常会导致人类神经精神疾病。