Regulation of Pet1/FEV binding and chromatin accessibility during serotonergic neuron development

项目总结/摘要 神经递质5-羟色胺（5-HT）与许多精神疾病的病理生理学有关， 神经发育障碍，包括焦虑、抑郁、自闭症、精神分裂症、注意力- 缺陷/多动障碍和强迫症。5-羟色胺神经元是如何成熟并获得它们的 传播者身份和成人特征仍然知之甚少。在胚胎期和早期 出生后，5-羟色胺神经元的谱系特异性基因表达模式由一个网络协调， 发育关键转录因子，包括ETS家族转录因子Pet 1（人 直系同源物FEV），其对于5-羟色胺神经传递的建立是必需的。有趣的是，我们最近 发现Pet 1在胎儿到出生后早期的过渡期转换目标，从控制5-HT的上调， 从胎儿时期的合成基因到出生后时期的突触兴奋性所需的激活基因。 本研究探讨了这一假设，即在染色质可及性的顺式调节的变化， 元件决定了转录过程中可用于Pet 1调控的转录靶点的库， 发展此外，我假设Pet 1结合也塑造了染色质结构，以指导细胞凋亡。 5-羟色胺神经元的基因表达轨迹。为了验证这些假设，在目标1中，我将绘制 5-羟色胺神经元的开放染色质全局分布 在多个胚胎和出生后早期发育时间进行高通量测序（ATAC-seq） 点，并探讨开放染色质与5-HT神经元基因表达的关系。在目标2中，我将分析 在与Aim 1相同的5-羟色胺神经元发育阶段期间Pet 1 DNA占有率的变化 使用Pet 1染色质免疫沉淀结合高通量测序（ChIP-seq），并确定 Pet 1对发育关键的染色质重塑或维持的重要性， 在Pet 1敲除小鼠中的seq。通过阐明Pet 1如何动态调节基因表达， 5-羟色胺神经元的成熟，我将提供深入了解神经精神疾病的病理生理学 5-羟色胺基因表达被认为受到干扰的情况。此外，这项研究提供了 宝贵的培训机会，使我能够熟练掌握小鼠遗传学和转录组学方面的技术， 表观遗传学分析，分子神经科学的广泛概念知识，以及实验研究的经验。 设计，数据解释，口头和书面沟通，这对我自己的成长至关重要， 正在接受培训的物理学家兼科学家。

PROJECT SUMMARY/ABSTRACT The neurotransmitter serotonin (5-HT) is implicated in the pathophysiology of many psychiatric and neurodevelopmental disorders, including anxiety, depression, autism, schizophrenia, attention- deficit/hyperactivity disorder, and compulsive disorders. How serotonin neurons mature and acquire their transmitter identity and adult characteristics remain poorly understood. During the embryonic and early postnatal period, lineage specific gene expression patterns of serotonin neurons are orchestrated by a network of developmentally critical transcription factors, including the ETS family transcription factor Pet1 (human ortholog FEV) that is essential for the establishment of serotonin neurotransmission. Intriguingly, we recently found Pet1 switches targets during fetal to early postnatal transition, from controlling the upregulation of 5-HT synthesis genes during fetal life to activating gene required for synaptic excitability during the postnatal period. This study investigates the hypothesis that the changes in the chromatin accessibility of cis-regulatory elements dictate the repertoire of transcriptional targets that are available for Pet1 regulation during development. Furthermore, I hypothesize that Pet1 binding also shapes chromatin architecture to direct the gene expression trajectories of developing serotonin neurons. To test these hypotheses, in Aim 1, I will map the global open chromatin landscape of serotonin neurons using Assay for Transposase Accessible Chromatin with high throughput sequencing (ATAC-seq) at multiple embryonic and early postnatal developmental time points, and investigate the relation of open chromatin to 5-HT neuron gene expression. In Aim 2, I will analyze the changes in Pet1 DNA occupancy during the same stages of serotonin neuron development as in Aim 1 using Pet1 chromatin immunoprecipitation coupled to high throughput sequencing (ChIP-seq), and determine the importance of Pet1 for developmentally critical chromatin remodeling or maintenance by performing ATAC- seq in Pet1 knockout mice. By elucidating how Pet1 dynamically regulates gene expression critical for the maturation of serotonin neurons, I will provide insight into the pathophysiology of the neuropsychiatric conditions in which serotonin gene expression is thought to be perturbed. Additionally, this study provides valuable training opportunity for me to gain technical proficiency in mouse genetics and transcriptomic and epigenetic profiling, broad conceptual knowledge in molecular neuroscience, and experience with experimental design, data interpretation, and oral and written communication that are crucial for my own growth as a physician-scientist in training.