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：
9344688
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/9344688
关键词：
18q Alleles Alpha Cell Animal Model Architecture Behavioral Bioinformatics Biological Models Biological Process Birth Brain CRISPR/Cas technology 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 Image Analysis Ion Channel Medial Mediating Mental disorders Modeling Molecular Molecular Analysis Molecular Biology Molecular Profiling Mus Mutation Neurodevelopmental Disorder Neurons Outcome Pathologic 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 design design and construction differential expression disorder risk experimental study foot gene function genetic risk factor in utero in vivo insight knock-down mouse model neurodevelopment neuronal excitability neuropsychiatric disorder novel novel therapeutics overexpression programs public health relevance relating to nervous system risk variant small hairpin RNA therapeutic development therapeutic target therapy design therapy development 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疾病（ASD）是慢性和衰弱的，病因学和病理生理学相对未知。最近的朝着了解这些疾病在人群层面的遗传结构的进展已导致识别许多遗传危险因素。但是，在大多数情况下，风险和鉴定基因的相关功能尚不清楚，因此可以识别治疗靶标仍然很困难。在我们的提议中，我们概述了确定治疗目标的路线图精神病障碍并提供了表明我们方法有价值的初步数据。我们建议使用两个模型系统，一个细胞自主模型，在该模型中，精神分裂症和ASD基因TCF4的表达（转录因子4）使用子宫电穿孔和具有组成型的小鼠模型来操纵模拟皮特 - 霍普金斯综合征（PTHS）的一个TCF4等位基因的种系截断。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和治疗的发展治疗将基于我们对负责分子机制的新兴理解。在AIM1中，我们假设TCF4在转录中调节了内在的神经元兴奋性，因此 TCF4表达的抑制作用将导致与PTH相关的神经元生理异常。我们的初步数据表明，在PFC的第2/3层锥体细胞中TCF4的子宫敲低中，异常的内在兴奋性和异位尖峰适应。我们显示了细胞机制这些表型与毕竟的过度过敏（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受候选离子通道的调节 TCF4和可能在PTH中观察到的认知表型的基础。为了验证这些靶基因，我们提出药理学救援和分子表拷贝实验。在AIM 2中，我们建议使用鼠标模型 Pths。我们的初步数据表明来自TCF4+/TR小鼠的PFC层2/3神经元显示出相似的内在性使用shRNA/crispr构建体敲除tcf4时，我们观察到的兴奋性缺陷。我们建议细胞和分子实验，以确定具有未来目标的这种表型的机制使用药理学来挽救这些小鼠的行为缺陷。在AIM 3中，我们建议确定在PTHS小鼠模型中，神经转录组在整个发育过程中发生了变化。我们假设识别 TCF4+/TR在神经转录组上的分子后果将提供机械和病理有关PTH和可能其他特发性ASD的洞察力。这些目标共同旨在确定治疗PTH的治疗靶标。