R-loops as Mechanisms Governing Neural Differentiation and Cell Type-Specific Transcription

R 环作为控制神经分化和细胞类型特异性转录的机制

• 批准号: 10118038
• 负责人: Elizabeth Ann LaMarca
• 金额: $ 2.95万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2021-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10118038
• 关键词: Affect; Brain; Brain region; Cells; ChIP-seq; Child; Complex; Cues; DNA; Data; Development; Developmental Gene; Diagnosis; Diagnostic; Disease; Epigenetic Process; Etiology; Expression Profiling; Functional disorder; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Goals; Human; Hybrids; Immunoprecipitation; In Vitro; Laboratory Research; Lead; Link; Maps; Measures; Mediating; Mental disorders; Messenger RNA; Modeling; Neurodevelopmental Disorder; Neuronal Differentiation; Neurons; Nuclear Pore Complex; Phenotype; Play; Population; Process; Public Health; RNA; RNA Polymerase II; Regulation; Research; Role; Schizophrenia; Synapses; Techniques; Testing; Time; Tissues; Training; Transcriptional Regulation; Treatment Efficacy; United States; Variant; autism spectrum disorder; base; brain cell; career; cell type; chromatin immunoprecipitation; developmental disease; diagnostic biomarker; differential expression; epigenomics; genome-wide; immunocytochemistry; improved; induced pluripotent stem cell; knock-down; multi-electrode arrays; nerve stem cell; neurodevelopment; neurogenesis; neuropsychiatric disorder; neuropsychiatry; new therapeutic target; next generation sequencing; novel; novel diagnostics; nucleic acid structure; overexpression; prenatal; relating to nervous system; research study; ribonuclease H1; synaptic function; synaptogenesis; therapeutic target; tool; transcriptome sequencing

PROJECT SUMMARY The neuropsychiatric disorders schizophrenia and autism together affect over 15% of children in the United States, representing a significant public health concern. Development of effective therapeutics and diagnostic tools for these disorders has been hindered by our incomplete understanding of their complex neurodevelopmental etiologies. Gene expression variation is a common attribute of these disorders, thought to arise from developmental stage- and tissue-specific RNA regulation during neurodevelopment. New genome- wide mapping strategies have identified a connection between R-loops (a three-stranded nucleic acid structure containing a DNA/RNA hybrid) and transcriptional regulation, suggesting a possible link between R-loops and gene expression variation. However, R-loops have never before been characterized on a genome-wide scale in the human brain, precluding research studying their role in neurodevelopmental illnesses. Our research has showed that R-loops may poise developmental genes for transcription during neural differentiation, and our immediate goal is to test this hypothesis by leveraging human induced pluripotent stem cell (hiPSC)-based models to functionally manipulate R-loop levels. Specifically, we aim to reduce R-loop levels in hiPSC-derived neural progenitor cells and measure the effect this has on transcription, differentiation, and ability of neural cells to form functional synapses. We aim to identify a causal mechanism of this R-loop-mediated transcriptional poising by exploring the hypothesis that R-loops stall RNA polymerase II, as stalled RNA polymerase II is thought to keep developmental genes transcriptionally inactive but ready for expression upon developmental cues. Importantly, we will map R-loops genome-wide for the first time in the developing human brain, and link R-loop distributional shifts during human neurogenesis with neurodevelopmental disease-associated gene expression signatures. Here, we will use cutting-edge techniques, including DNA/RNA immunoprecipitation followed by next- generation sequencing, to achieve these goals. These first steps toward understanding R-loop function in neural cells will facilitate our long-term objective to uncover the epigenetic mechanisms of aberrant RNA regulation in neuropsychiatric illness, in order to improve diagnostics and identify novel therapeutic targets for these illnesses.

项目概要 神经精神疾病精神分裂症和自闭症共同影响了超过 15% 的儿童 美国，代表着重大的公共卫生问题。开发有效的治疗方法和 由于我们对其复杂性的不完全了解，这些疾病的诊断工具受到阻碍 神经发育病因学。基因表达变异是这些疾病的共同属性，被认为是 由神经发育过程中发育阶段和组织特异性 RNA 调节产生。新基因组- 广泛的作图策略已经确定了 R 环（三链核酸结构 包含 DNA/RNA 杂合体）和转录调控，表明 R 环和转录调控之间可能存在联系 基因表达变异。然而，R 环以前从未在全基因组范围内被表征过。 人类大脑，排除了研究它们在神经发育疾病中的作用的研究。我们的研究有 研究表明，R 环可能会在神经分化过程中平衡发育基因的转录，并且我们的研究 近期目标是利用基于人类诱导多能干细胞 (hiPSC) 的技术来检验这一假设 模型来功能性地操纵 R 环水平。具体来说，我们的目标是降低 hiPSC 衍生的 R 环水平 神经祖细胞并测量其对神经细胞转录、分化和能力的影响 形成功能性突触。我们的目标是确定这种 R 环介导的转录的因果机制 通过探索 R 环使 RNA 聚合酶 II 失速的假设，正如人们认为的那样，R 环使 RNA 聚合酶 II 失速 保持发育基因转录失活，但准备好根据发育线索表达。 重要的是，我们将首次在发育中的人类大脑中绘制全基因组范围的 R 环图谱，并将 R 环连接起来 人类神经发生过程中神经发育疾病相关基因表达的分布变化 签名。在这里，我们将使用尖端技术，包括 DNA/RNA 免疫沉淀，然后是- 世代测序，以实现这些目标。理解神经网络中 R 环函数的第一步 细胞将促进我们的长期目标，即揭示异常 RNA 调控的表观遗传机制 神经精神疾病，以改进诊断并确定这些疾病的新治疗靶点。