Gene regulatory mechanisms controlling development of serotonin neuron subtypes

虽然在了解遗传起源方面取得了重大进展， 神经发育障碍，目前尚不清楚哪些特定的分子步骤被破坏，以及在哪些 特定的神经元类型，它们功能失调或无法操作。这种缺乏理解的一个潜在原因是 神经元最初是根据产生的递质类型分类的，现在众所周知， 实质性的分子、细胞和功能异质性。似乎神经发育 疾病不仅可能源于一个特定群体的整个群体的发育改变的身份， 神经元类型，但也从其特定的分子或功能亚型之一的发育改变。那里 几十年来，人们一直对控制5-HT神经元的调节机制感兴趣，因为5-HT具有 对中枢神经回路和多巴胺能系统功能障碍的广泛调节作用， 与几种神经精神疾病有关，包括抑郁症，与压力有关的焦虑症， 自闭症、智力障碍、强迫症和精神分裂症。5-HT神经元具有巨大的分子， 形态和电生理异质性。然而，5-HT神经元的发育轨迹 亚型目前尚不清楚，控制其发展的调节机制也是未知的。的 本研究的目的是利用单细胞RNA-seq和单细胞ATAC-seq， 确定5-HT神经元亚型转录组和染色质的时空发育轨迹 可访问性。我们将联合收割机结合单细胞基因组学方法的最新进展， 建立了多巴胺能转基因工具，我们在流式分选小鼠5-HT神经元方面的丰富经验， 我们的生物信息学专业知识，研究单细胞5-HT神经元转录组的发展， 从胎儿到出生后早期成熟的染色质可及性。我们还将在单个细胞中进行研究 水平，假设5-HT神经元中的两个疾病相关的终末选择因子，Pet 1和Lmx 1b， 通过亚型特异性基因的差异调节决定有丝分裂后5-HT神经元亚型 表达、亚型特异性染色质可及性和下游亚型特异性转录的控制 因子码Pet 1作为FEV中的纯合敲除突变特别令人感兴趣，FEV是 Pet 1，最近在两个患有智力残疾和自闭症谱系障碍的兄弟中报道。在Aim中 1，我们将定义5-HT神经元亚型的发育轨迹。在目标2中，我们将研究 Pet 1和Lmx 1b控制5-HT神经元亚型转录组。在目标3中，我们将确定染色质 参与5-HT神经元亚型生成的机制。实现我们提出的目标， 导致对产生5-HT神经元的亚型特异性基因调控网络的更深入理解 亚型，这可能有助于阐明在神经发育过程中被破坏的特定神经遗传通路。 例如ASD/ID。

Although significant progress has been made in understanding the genetic origins of neurodevelopmental disorders, it remains unclear what specific molecular steps are disrupted and in which specific neurons types they are dysfunctional or inoperable. One potential reason for this lack of understanding is that neurons, originally classified according the type of transmitter produced, are now well known to possess substantial molecular, cellular, and functional heterogeneity. It seems plausible that neurodevelopmental disorders may arise not only from developmentally altered identities of an entire population of one particular neuron type but also from altered development of one of its specific molecular or functional subtype(s). There has been a decades-long intense interest in the regulatory mechanisms controlling 5-HT neurons as 5-HT has wide-ranging modulatory effects on central neural circuitry and dysfunction of the serotonergic system has been implicated in several neuropsychiatric diseases including depression, stress-related anxiety disorders, autism, intellectual disability, OCD, and schizophrenia. 5-HT neurons possess tremendous molecular, morphological, and electrophysiological heterogeneity. However, developmental trajectories of 5-HT neuron subtypes are currently unknown as are the regulatory mechanisms that govern their development. The objective of the proposed research is to use single cell RNA-seq and single cell ATAC-seq to comprehensively define the spatiotemporal developmental trajectories of 5-HT neuron subtype transcriptomes and chromatin accessibility. We will combine recent advances in single-cell genomics methods together with our well- established serotonergic transgenic tools, our extensive experience in flow sorting mouse 5-HT neurons, and our bioinformatics expertise to investigate the development of single-cell 5-HT neuron transcriptomes and chromatin accessibility throughout fetal to early postnatal maturation. We will also investigate at the single cell level, the hypothesis that the two disease-associated terminal selectors in 5-HT neurons, Pet1 and Lmx1b, function to determine postmitotic 5-HT neuron subtypes through differential regulation of subtype-specific gene expression, subtype-specific chromatin accessibility and control of downstream subtype-specific transcription factor codes. Pet1 is of special interest as homozygous knockout mutations in FEV, the human ortholog of Pet1, were recently reported in two brothers with Intellectual Disability and Autism Spectrum Disorder. In Aim 1, we will define the developmental trajectories of 5-HT neuron subtypes. In Aim 2, we will investigate the control of 5-HT neuron subtype transcriptomes by Pet1 and Lmx1b. In Aim 3, we will determine the chromatin mechanisms involved in the generation of 5-HT neuron subtypes. The completion of our proposed aims will lead to a greater understanding of the subtype-specific gene regulatory networks that generate 5-HT neuron subtypes, which may help illuminate specific neurogenetic pathways that are disrupted in neurodevelopmental disorders such as ASD/ID.