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系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它们的活性，或者通过Cre介导的基因敲除来评估基因功能。大多数Cre驱动系，如基于BAC的Cre驱动或Cre敲入特定基因座，监测整个遗传基因座的完整表达模式。然而，很少有基因专门表达在特定神经元类型的非常小的群体中，并且这种细胞特异性的缺乏限制了这些驱动系的使用。我们建议在这里开发转基因小鼠驱动线，指导Cre表达的神经元细胞类型在小鼠大脑的不同区域的数量非常有限，从而提供工具，以精确地映射其功能和分子组成。为了实现这一目标，我们的目标是测试假设-从我们过去在线虫中的工作中建立-小鼠基因座的顺式调节控制元件编码脊椎动物中枢神经系统中四种主要神经递质系统的囊泡转运蛋白，谷氨酸和-氨基丁酸（GABA）和乙酰胆碱（ACh）是由个体的模块化组装组成的，高度细胞类型特异性顺式调节元件。我们将实验测试的假设，表达的个人，孤立的顺式调节元件可能细分胆碱能，谷氨酸能和GABA能域到有限的，也许是新的领域的小鼠中枢神经系统，从而构成可重复的和高度特异性的驱动程序，指导基因的表达，允许遗传操纵的神经元和神经元电路。这种顺式调节切割方法可以解决大多数目前可用的驱动线的特异性问题，这些驱动线不能专门靶向有限数量的细胞。