Molecular and Behavioral Neurobiology of Transcription Factor TCF4

描述(申请人提供)：基本螺旋-环-螺旋转录因子TCF4(又名E2-2)广泛参与人类中枢神经系统功能。TCF4在几个大型的GWAS研究中被证实是罕见的高度重复的精神分裂症易感基因之一，当TCF4缺乏时是皮特-霍普金斯综合征(PTHS)的致病因素。PTHS在人类认知功能方面具有令人信服的特征，与明显的记忆缺陷、自闭症行为有关，更重要的是，几乎完全缺乏语言发展。因此，了解TCF4在中枢神经系统中的作用和功能对于人类的语言和听觉认知、记忆功能、自闭症谱系行为和精神分裂症的易感性具有重要意义。尽管TCF4在人类中枢神经系统中的功能非常重要，但对TCF4的基础神经生物学研究还很少。因此，本项目将使用基因工程小鼠模型来评估TCF4在记忆、社会互动和交流、海马突触可塑性、突触解剖以及中枢神经系统的表观基因组和转录调控中的作用。拟议研究的中心假设是，TCF4通过主动调节转录活动来调节大脑触发长期突触可塑性和记忆形成的能力，以响应行为经验。该方法将使用胚系结构性敲除和成年小鼠海马区TCF4的急性敲除或缺失，结合广泛的行为、电生理和转录/表观基因组学特征，来确定TCF4功能是否对成熟中枢神经系统的正常记忆和突触可塑性是持续的必需的。这些研究不仅将利用现有的生殖系基因敲除系，还将利用已有的TCF4等位基因小鼠系，并将这些小鼠与成年海马区病毒驱动的cre表达和发育后CaMKII启动子驱动的前脑神经元选择性cre表达相结合，以实现体内CNS TCF4功能的可诱导的发育后衰减。使用这些新颖的鼠标模型，我们将实现三个具体目标。目的1验证TCF4缺陷小鼠表现出认知记忆和社会互动缺陷的假说，以及TCF4缺陷小鼠突触可塑性改变的假说。目标2将包括一个重要的翻译 并将检验转录促进组蛋白脱乙酰酶抑制物(HDACi)将恢复TCF4缺陷小鼠的学习、记忆和突触可塑性的假设，作为PTHS模型中行为缺陷和可塑性缺陷的概念证明 HDACi可以从药理上逆转小鼠的病情。目的3将验证TCF4功能丧失触发成熟中枢神经系统染色质结构、DNA甲基化和基因转录的继发性改变的假设。为此，我们将使用无偏见的方法，利用MBD-seq、mRNA-seq和小RNA-seq方法来全面分析TCF4缺陷小鼠潜在的转录和表观基因组变化。

DESCRIPTION (provided by applicant): The basic helix-loop-helix transcription factor TCF4 (aka E2-2) has been implicated broadly in human CNS function. TCF4 was confirmed in several large GWAS studies as one of the rare highly replicated schizophrenia susceptibility genes, and when haplo-insufficient TCF4 is the causative factor for Pitt-Hopkins Syndrome (PTHS). PTHS has compelling attributes in terms of human cognitive function, being associated with pronounced memory deficits, autistic behaviors, and importantly an almost complete lack of language development. Thus, understanding the roles and function of TCF4 in the CNS is highly significant with respect to human language and auditory cognition, memory function, autism spectrum behavior, and schizophrenia susceptibility. Despite the clear importance of TCF4 function in the human CNS, the basic neurobiology of TCF4 has been only sparsely studied. Thus, this Project will use genetically engineered mouse models to assess the role of TCF4 in memory, social interactions and communication, hippocampal synaptic plasticity, synaptic anatomy, and epigenomic and transcriptional regulation in the CNS. The central hypothesis of the proposed studies is that TCF4 regulates the brain's ability to trigger long-term synaptic plasticity and memory formation by actively regulating transcriptional activity in response to behavioral experience. The approach will be to use both germline constitutive knockout and acute knockdown or deletion of TCF4 in the adult mouse hippocampus, coupled with extensive behavioral, electrophysiological, and transcriptomic/epigenomic characterization, to determine if TCF4 function is necessary in an ongoing fashion for normal memory and synaptic plasticity in the mature CNS. The studies will capitalize not only upon the available germline knockout line but also upon an already-available floxed TCF4 allele mouse line, and combine these mice with both virus- driven cre expression in the adult hippocampus and post-developmental CaMKII promoter-driven forebrain neuron-selective cre expression to achieve inducible post-developmental attenuation of CNS TCF4 function in vivo. Using these novel mouse models we will undertake three Specific Aims. Aim 1 will test the hypothesis that TCF4-deficient mice exhibit cognitive memory and social interaction deficits, and also test the hypothesis that TCF4-deficient mice exhibit altered synaptic plasticity. Aim 2 will comprise a significant translational component of the studies and will test the hypothesis that transcription-promoting Histone DeAcetylase Inhibitors (HDACi) will restore learning, memory, and synaptic plasticity in TCF4-deficient mice, as proof-of- concept that behavioral deficits and plasticity deficits in PTHS model mice can be reversed pharmacologically with HDACi. Aim 3 will test the hypothesis that loss of TCF4 function triggers secondary alterations in chromatin structure, DNA methylation, and gene transcription in the mature CNS. Toward this end, we will use the unbiased approach of utilizing MBD-seq, mRNA-seq, and small RNA-seq approaches for a comprehensive analysis of potential transcriptomic and epigenomic alterations in TCF4-deficient mice.