Cellular and Molecular Analysis of the Schizophrenia and Autism Spectrum Disorder gene Transcription Factor 4 (TCF4)

精神分裂症和自闭症谱系障碍基因转录因子 4 (TCF4) 的细胞和分子分析

基本信息

批准号：
9158209
负责人：
BRADY J MAHER
金额：
$ 45.65万
依托单位：
LIEBER INSTITUTE, INC.
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-02 至 2020-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9158209
关键词：
Alleles Animal Model Architecture Behavioral Bioinformatics Biological Models Biological Process Birth Brain Cell model Cells Cellular biology Chronic Clinical Cognitive Cognitive deficits Complex Data Development Disease Electrophysiology (science)Electroporation Etiology Experimental Designs Frequencies Functional disorder Future Gene Expression Gene Targeting Genes Genetic Genetic Risk Genetic Transcription Genotype Goals Image Ion Channel Medial Mediating Mental disorders Modeling Molecular Molecular Analysis Molecular Biology Molecular Profiling Mus Mutation Neurodevelopmental Disorder Neurons Outcome Pathway interactions Pharmacology Phenocopy Phenotype Physiological Physiology Platelet Factor 4 Population Prefrontal Cortex Process Proteins Public Health Pyramidal Cells Rattus Research Risk Rodent Schizophrenia Slice Syndrome Techniques Time Transcript autism spectrum disorder base cell type chromatin immunoprecipitation chromosome 18q deletion syndrome design design and construction differential expression foot genetic risk factor in utero in vivo insight knock-down molecular imaging mouse model neurodevelopment neuronal excitability neuropsychiatric disorder novel novel therapeutics overexpression programs public health relevance relating to nervous system research study risk variant small hairpin RNA therapeutic development therapeutic target therapy design transcription factor transcriptome transcriptome sequencing treatment strategy

项目摘要

PROJECT SUMMARY: Neurodevelopmental disorders including Schizophrenia and Autism spectrum disorders (ASD) 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 propose to use two model systems, a cell autonomous model in which expression of the schizophrenia and ASD gene TCF4 (transcription factor 4) is manipulated using in utero electroporation and a mouse model that has a constitutive germline truncation of one TCF4 allele that models Pitt-Hopkins syndrome (PTHS). We will characterize the resulting neuronal phenotypes using brain slice electrophysiology, cell biology, and confocal imaging. Identified 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. In Aim1, we hypothesize that TCF4 transcriptionally regulates intrinsic neuronal excitability and therefore suppression of TCF4 expression will result in abnormal neuronal physiology relevant to PTHS. Our preliminary data suggest that in utero knockdown of TCF4 in layer 2/3 pyramidal cells of the PFC results in abnormal intrinsic excitability and ectopic spike-frequency adaptation. We show the cellular mechanisms of these phenotypes are associated with an increase in the afterhyperpolarization (AHP). Using a novel molecular profiling technique (iTRAP) we have identified two candidate ion channels that are regulated by TCF4 and may underlie cognitive phenotypes observed in PTHS. To validate these target genes, we propose pharmacological rescue and molecular phenocopy experiments. In Aim 2, we propose to use a mouse model of PTHS. Our preliminary data indicates PFC layer 2/3 neurons from TCF4+/tr mice show similar intrinsic excitability deficits to what we observe when TCF4 is knockdown using shRNA/Crispr constructs. We propose cellular and molecular experiments to identify the mechanisms underlying this phenotype with the future goal of using pharmacology to rescue behavioral deficits in these mice. In Aim 3, we propose to identify how the neural transcriptome is altered across development in PTHS mouse model. We hypothesize that identifying molecular consequences of TCF4+/tr on the neural transcriptome will provide mechanistic and pathological insight about PTHS and potentially other idiopathic ASDs. Together, these Aims are designed to identify therapeutic targets for treatment of PTHS.

项目摘要：神经发育障碍，包括精神分裂症和自闭症谱系自闭症谱系障碍 (ASD) 是一种慢性且使人衰弱的疾病，其病因和病理生理学相对未知。最近的在人口水平上了解这些疾病的遗传结构方面取得的进展导致识别许多遗传风险因素。然而，在大多数情况下，风险和风险的分子机制所识别基因的相关功能尚不清楚，因此需要确定治疗靶点仍然很困难。在我们的提案中，我们概述了确定治疗靶点的路线图精神疾病并提供初步数据表明我们的方法是有价值的。我们建议使用两个模型系统，一个细胞自主模型，其中表达精神分裂症和 ASD 基因 TCF4 （转录因子 4）使用子宫内电穿孔和具有组成型的小鼠模型进行操作一种模拟皮特-霍普金斯综合征 (PTHS) 的 TCF4 等位基因的种系截短。我们将表征使用脑切片电生理学、细胞生物学和共聚焦成像产生的神经元表型。确定的表型将被评估为潜在的病理生理学和治疗的开发治疗将基于我们对相关分子机制的新认识。在 Aim1 中，我们假设 TCF4 转录调节内在神经元兴奋性，因此 TCF4表达的抑制将导致与PTHS相关的神经元生理学异常。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。我们展示了细胞机制这些表型与后超极化（AHP）的增加有关。使用小说分子分析技术（iTRAP）我们已经确定了两个候选离子通道，它们受 TCF4 可能是 PTHS 中观察到的认知表型的基础。为了验证这些目标基因，我们建议药理学救援和分子表型实验。在目标 2 中，我们建议使用小鼠模型 PTHS 的。我们的初步数据表明 TCF4+/tr 小鼠的 PFC 层 2/3 神经元表现出相似的内在特征当使用 shRNA/Crispr 构建体敲低 TCF4 时，我们观察到兴奋性缺陷。我们建议细胞和分子实验以确定该表型背后的机制以及未来的目标使用药理学来挽救这些小鼠的行为缺陷。在目标 3 中，我们建议确定如何 PTHS 小鼠模型的神经转录组在发育过程中发生变化。我们假设识别 TCF4+/tr 对神经转录组的分子影响将提供机制和病理学关于 PTHS 和潜在的其他特发性 ASD 的见解。这些目标共同旨在确定 PTHS 治疗的治疗目标。