Developing drivers for neuron type-specific gene expression

开发神经元类型特异性基因表达的驱动程序

• 批准号: 8935927
• 负责人: Oliver Hobert
• 金额: $ 62.6万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 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/8935927
• 关键词: Acetylcholine; Adult; Aminobutyric Acids; Binding Sites; Brain; Caenorhabditis elegans; Cell Count; Cells; DNA; Dissection; Elements; Enzymes; Future; Gene Expression; Generations; Genes; Genetic; Genomics; Glutamates; Health; Individual; Integrase; LacZ Genes; Maps; Mediating; Molecular; Monitor; Mus; Mutate; Nature; Nematoda; Nervous system structure; Neuraxis; Neurons; Neurosciences; Neurotransmitters; Nucleic Acid Regulatory Sequences; Pattern; Phenotype; Population; Positioning Attribute; Proteins; Regulatory Element; Reporter; Specificity; System; Testing; Transgenic Mice; Transgenic Organisms; Work; acetylcholine transporter; base; cell type; cholinergic; gene function; genetic manipulation; insight; interest; knockout gene; mouse genome; neuronal circuitry; novel; optogenetics; theories; tool; transcription factor

 DESCRIPTION (provided by applicant): Driver lines that direct Cre protein to specific neuron types have proven to be invaluable tools to not only visualize specific neuron types but also to manipulate their activity through the Cre- mediated activation of optogenetic probes or to assess gene function by Cre-mediated gene knockout. Most Cre driver lines, such as BAC-based Cre drivers or knock-ins of Cre into specific loci, monitor the complete expression pattern of entire genetic loci. However, very few genes are exclusively expressed in very small populations of specific neuron types and this lack of cellular specificity limits the use of these driver lines. W propose here to develop transgenic mouse driver lines that direct Cre expression to very restricted numbers of neuronal cell types in different regions of the mouse brain, thereby providing tools to precisely map their function and molecular composition. To achieve this aim, we aim to test the hypothesis - built from our past work in the nematode C.elegans - that the cis-regulatory control elements of the mouse loci that encode the vesicular transporters for the four main neurotransmitter systems in the vertebrate central nervous system, glutamate and -aminobutyric acid (GABA) and acetylcholine (ACh) are composed of a modular assembly of individual, highly cell type-specific cis-regulatory elements. We will experimentally test the hypothesis that the expression of individual, isolated cis-regulatory elements may subdivide cholinergic, glutamatergic and GABAergic domains into restricted and perhaps novel domains of the mouse central nervous system and thereby constitute reproducible and highly specific drivers for directing the expression of genes that allow the genetic manipulation of neurons and neuronal circuits. This cis-regulatory dissection approach may solve the specificity problem of most currently available driver lines that are unable to exclusively target restricted numbers of cells.

 描述（由申请人提供）：将 Cre 蛋白导向特定神经元类型的驱动线已被证明是非常宝贵的工具，不仅可以可视化特定神经元类型，还可以通过 Cre 介导的光遗传学探针激活来操纵其活性或评估基因功能大多数 Cre 驱动程序系（例如基于 BAC 的 Cre 驱动程序或将 Cre 敲入特定位点）可监测整个基因位点的完整表达模式，但是，很少有基因专门在非常小的基因座中表达。人口为特定的神经元类型和细胞特异性的缺乏限制了这些驱动细胞系的使用，W 在此建议开发转基因小鼠驱动细胞系，将 Cre 表达引导至小鼠大脑不同区域中数量非常有限的神经元细胞类型，从而为精确绘制它们的功能和分子组成为了实现这一目标，我们的目标是测试基于我们过去在线虫研究中建立的假设，即编码囊泡转运蛋白的小鼠基因座的顺式调控元件。四个主要脊椎动物中枢神经系统中的神经递质系统，谷氨酸和氨基丁酸（GABA）和乙酰胆碱（ACh）是由单个细胞类型特异性顺式调节元件的模块化组装组成的，我们将通过实验高度检验表达的假设。单个的、分离的顺式调节元件可以将胆碱能、谷氨酸能和GABA能结构域细分为小鼠中枢神经系统的限制性的和可能是新颖的结构域，从而构成可重复且高度特异性的驱动程序，用于指导基因的表达，从而允许对神经元和神经元回路进行遗传操作。这种顺式调节解剖方法可以解决大多数当前可用的驱动程序系无法专门针对有限数量的细胞的特异性问题。 。