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Identification of enhancers whose activity defines cortical interneuron types

识别其活性定义皮质中间神经元类型的增强子

基本信息

批准号：
8935930
负责人：
JOHN L. R. RUBENSTEIN
金额：
$ 48.83万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-09-26 至 2017-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8935930
关键词：
Adult Agreement Alleles Autistic Disorder Automobile Driving Behavioral Binding Biological Assay Brain Cell Transplants Cells Cerebral cortex ChIP-seq Development Disease Electrophysiology (science)Embryo Enhancers Epilepsy Funding Gene Expression Genes Genetic Enhancer Element Genetic Transcription Genomics Health Histones Human Image Immunohistochemistry In Situ Hybridization Interneuron function Interneurons Knowledge Label Lentivirus Vector Maps Medial Methods Molecular Molecular Genetics Morphology Mus Nature Neonatal Neurons Neurophysiology - biologic function Pattern Phenotype Physiological Physiology Play Positioning Attribute Process Property Proteins RNA Regulatory Element Reporter Research Rhodopsin Role Schizophrenia Series Slice Staging Taxonomy Time Transplantation Viral Whole-Cell Recordings base behavior measurement behavioral study cell type in vivo neurogenetics neuropsychiatry novel optogenetics progenitor ranpirnase research study tool transcription factor

项目摘要

DESCRIPTION (provided by applicant): Molecular definitions of neural cell types largely depend on the expression of RNAs or proteins as assessed by in situ hybridization, RNA array and sequencing, and immunohistochemistry. However, recent studies are demonstrating that gene regulatory elements, such as enhancers, can have highly specific spatial and temporal activity patterns in the developing brain. Thus, enhancer activity can be used to define neural cell types, and importantly, also have other broad applications. First, they can be used as tools to drive gene expression in specific cell types, which can then be used to visualize and/or purify the cells (GFP), modify gene expression in the cells (Cre), modify electrical activity (channel rhodopsin), and visualize electrical activity in the cells (GCaMP). Secondly, knowledge about the nature and position of enhancers enables geneticists to identify disease alleles that map in extra-exonic genomic space. Herein, we will focus on identifying enhancers that are active in cortical interneurons, during development and in the mature state. We choose to study cortical GABAergic interneurons because of their central role in cortical function and diseases. We will focus on these GABAergic neurons derived from the medial ganglionic eminence (MGE), which generates the majority of cortical interneurons. Our aim is to identify novel enhancers that are active in cortical interneurons using three assays: 1) histone (H3K27Ac) ChIP-seq (marker of active enhancers); 2) transcription factor (TF) ChIP- seq using TFs that regulate cortical interneurons development and function (Arx, Dlx2 and Lhx6); 3) a newly developed in vivo assay of enhancer function based on viral transduction of the enhancer driving Cre and GFP in immature cortical interneurons. We will then use a series of methods to define the interneuron subtypes that have enhancer activity. Together, our unique approach should define interneuronal cell types (developmental and adult), and simultaneously generate a powerful toolkit that will enable new ways to assess neural function and disease.

描述(申请人提供)：神经细胞类型的分子定义在很大程度上取决于通过原位杂交、RNA阵列和测序以及免疫组织化学评估的RNA或蛋白质的表达。然而，最近的研究表明，基因调控元件，如增强剂，可以在发育中的大脑中具有高度特定的空间和时间活动模式。因此，增强子活性可用于定义神经细胞类型，重要的是，还具有其他广泛的应用。首先，它们可以被用作在特定细胞类型中驱动基因表达的工具，然后可以用于可视化和/或纯化细胞(GFP)、修改细胞中的基因表达(Cre)、修改电活动(通道视紫红质)和可视化细胞中的电活动(GCaMP)。其次，关于增强子的性质和位置的知识使遗传学家能够识别在外显子基因组空间中映射的疾病等位基因。在这里，我们将专注于鉴定在皮质中间神经元中活跃的增强子，在发育过程中和成熟状态。我们选择研究皮质GABA能中间神经元，因为它们在皮质功能和疾病中起着核心作用。我们将关注这些来源于内侧神经节隆起(MGE)的GABA能神经元，MGE产生大部分皮质中间神经元。我们的目标是通过三种分析方法来确定在皮质间神经元中活跃的新的增强子：1)组蛋白(H3K27Ac)CHIP-SEQ(活性增强子的标记)；2)转录因子(TF)CHIP-SEQ，使用调节皮质间神经元发育和功能的转录因子(Arx、Dlx2和Lhx6)；3)基于病毒转导驱动未成熟皮质间神经元CRE和GFP的增强子的体内实验。然后，我们将使用一系列方法来定义具有增强活性的中间神经元亚型。总之，我们独特的方法应该定义神经元间细胞类型(发育和成人)，并同时产生一个强大的工具包，将使新的方法能够评估神经功能和疾病。