Functional analysis of the Schizophrenia and Autism Spectrum Disorder gene TCF4 i

精神分裂症和自闭症谱系障碍基因 TCF4 i 的功能分析

基本信息

批准号：
8889789
负责人：
BRADY J MAHER
金额：
$ 45.75万
依托单位：
LIEBER INSTITUTE, INC.
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：
2014-08-15 至 2016-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8889789
关键词：
Acute Architecture Autistic Disorder Autopsy Biological Brain Cell model Cellular Assay Cellular biology Chronic Clinical Complex Cortical Column Data Defect Development Diagnosis Disease Disease model Down-Regulation Electrophysiology (science)Electroporation Ephrins Etiology Exons Frequencies Functional disorder Genes Genetic Genetic Risk Genetic Transcription Goals Health Image Immigration In Vitro Individual Ion Channel Medial Mental disorders Modeling Molecular Neurodevelopmental Disorder Neurons Outcome Pathogenesis Pathway interactions Patients Pattern Phenocopy Phenotype Physiological Physiology Platelet Factor 4 Population Prefrontal Cortex Process Protein Isoforms Proteins Public Health Pyramidal Cells RNA Sequences RNA Splicing Radial Rattus Recombinants Research Risk Rodent Rodent Model Schizophrenia Signal Transduction Slice Syndrome System Techniques Testing Transcript Variant Wit autism spectrum disorder base chromosome 18q deletion syndrome data modeling gene function genetic association genetic risk factor genome wide association study human RNA sequencing in utero in vivo innovation insight migration molecular imaging neurodevelopment neuron development novel novel therapeutics programs research study risk variant sensor therapeutic development therapeutic target therapy design transcription factor treatment strategy

项目摘要

DESCRIPTION (provided by applicant): Neurodevelopmental disorders including Schizophrenia and Autism are chronic and debilitating, with relatively unknown etiology and pathophysiology. Recent progress towards understanding the genetic architecture of these disorders at the population level has led to the identification of many genetic risk factors. However, in most cases the molecular mechanism of risk and the relevant functions of the identified genes are not known and therefore identifying therapeutic targets remains difficult. In our proposal we have outlined a roadmap for the identification of therapeutic targets for psychiatric disorders and provide preliminary data that suggests our approach has merit. We will start by creating a testable cellular model of risk by genetically manipulating the expression of the schizophrenia and autism spectrum disorder gene TCF4 using in utero electroporation of the developing rat prefrontal cortex (PFC). We will then characterize the resulting neuronal phenotypes using acute brain slice electrophysiology, cell biology, and Ca2+ imaging. Such phenotypes will be evaluated as potentially pathophysiological and the development of therapeutic treatments will be based on our emerging understanding of the molecular mechanism responsible and by validating these mechanisms with molecular and pharmacological rescue or phenocopying. In Aim1, we hypothesize that TCF4 transcriptionally regulates the expression of ion channels genes that are necessary for normal neuronal physiology and in particular channels and/or Ca2+ sensors that underlie the afterhyperpolarization (AHP). Our preliminary data suggest that in utero knockdown of TCF4 in PFC layer 2/3 pyramidal cells results in abnormal intrinsic excitability and ectopic spike-frequency adaptation. We show the mechanisms of these phenotypes are associated with an increase in the AHP and are rescued by decreasing Ca2+ influx. We propose experiments to more specifically identify the mechanisms responsible. In Aim 2, we show that over-expression of TCF4 results in neuronal migration defects and the formation of abnormal cortical microcolumns in the developing PFC. We provide preliminary data that suggests these phenotypes may involve abnormal Eph/ephrin signaling and we hypothesize that accelerated neuronal migration will augment the development of intrinsic excitability and consequently disrupt the neuron's ability to integrate into the surrounding circuit. In Aim 3, we provide novel RNA sequencing data from postmortem brains of schizophrenia patients and controls that identifies a specific 5' exon of TCF4 that is differentially expressed by diagnosis, associated wit GWAS positive SNPs in TCF4, and is unique to a single TCF4 isoform (TCF4H). We propose to apply this new information to our roadmap by altering TCF4H expression in utero to more effectively model schizophrenia risk and provide additional validity to cellular models. We think these innovative approaches will enable us to provide significant insights into the etiology and pathophysiology of these disorders and will ultimately open doors to novel therapeutic treatments.

描述（由申请人提供）：包括精神分裂症和自闭症在内的神经发育障碍是慢性且使人衰弱的，其病因和病理生理学相对未知。最近在人群水平上了解这些疾病的遗传结构方面取得的进展导致了许多遗传风险因素的识别。然而，在大多数情况下，风险的分子机制和已识别基因的相关功能尚不清楚，因此确定治疗靶点仍然很困难。在我们的提案中，我们概述了确定精神疾病治疗靶点的路线图，并提供了表明我们的方法有价值的初步数据。我们将首先使用发育中的大鼠前额皮质 (PFC) 的子宫内电穿孔来基因操纵精神分裂症和自闭症谱系障碍基因 TCF4 的表达，从而创建可测试的风险细胞模型。然后，我们将使用急性脑切片电生理学、细胞生物学和 Ca2+ 成像来表征由此产生的神经元表型。此类表型将被评估为潜在的病理生理学，并且治疗方法的开发将基于我们对相关分子机制的新兴理解，并通过分子和药理学救援或表型复制来验证这些机制。在 Aim1 中，我们假设 TCF4 转录调节正常神经元生理学必需的离子通道基因的表达，特别是后超极化 (AHP) 基础的通道和/或 Ca2+ 传感器。我们的初步数据表明，在子宫内敲低 PFC 2/3 层锥体细胞中的 TCF4 会导致异常的内在兴奋性和异位尖峰频率适应。我们证明这些表型的机制与 AHP 的增加有关，并通过减少 Ca2+ 流入来挽救。我们提出实验来更具体地确定负责的机制。在目标 2 中，我们发现 TCF4 的过度表达会导致神经元迁移缺陷，并在发育中的 PFC 中形成异常皮质微柱。我们提供的初步数据表明这些表型可能涉及异常的 Eph/ephrin 信号传导，并且我们假设加速的神经元迁移将增强内在兴奋性的发展，从而破坏神经元整合到周围回路的能力。在目标 3 中，我们提供了精神分裂症患者和对照死后大脑的新 RNA 测序数据，该数据识别了 TCF4 的特定 5' 外显子，该外显子在诊断时差异表达，与 TCF4 中的 GWAS 阳性 SNP 相关，并且是单个 TCF4 亚型所独有的（TCF4H）。我们建议通过改变子宫内 TCF4H 的表达，将这一新信息应用到我们的路线图中，以更有效地模拟精神分裂症风险，并为细胞模型提供额外的有效性。我们认为这些创新方法将使我们能够对这些疾病的病因学和病理生理学提供重要的见解，并最终为新的治疗方法打开大门。