Epigenetic regulation of neural stem cell biology by Tet DNA dioxygenases

Tet DNA 双加氧酶对神经干细胞生物学的表观遗传调控

基本信息

批准号：
10386217
负责人：
Ian Campbell MacArthur
金额：
$ 5.1万
依托单位：
ALBERT EINSTEIN COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-09 至 2026-02-08
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10386217
关键词：
4-Hydroxy-Tamoxifen Apoptosis Astrocytes Biology Brain Carbon Cell Line Cell Maintenance Cell Survival Cerebral cortex Craniofacial Abnormalities Cytosine DNA DNA Methylation DNA Modification Process DNA biosynthesis Data Defect Development Dioxygenases Disease Down-Regulation Embryo Embryonic Development Enterobacteria phage P1 Cre recombinase Enzymes Epigenetic Process Etiology Excision Exhibits Failure Family Gene Expression Gene Expression Regulation Genes Genetic Transcription Goals Histones Human Intellectual functioning disability Joints Knock-out Knowledge Link LoxP-flanked allele Maintenance Maps Mediating Mentorship Methylation Methyltransferase Modeling Modification Mus Nerve Degeneration Nervous system structure Neuroanatomy Neurodevelopmental Disorder Neurons Oligodendroglia Oxides Phenotype Physicians Play Positioning Attribute Prosencephalon Proteins Recurrence Regulator Genes Regulatory Element Reporting Research Research Training Role Scientist Syndrome Tamoxifen Testing Tetanus Helper Peptide Tissues Transgenes bisulfite sequencing brain tissue craniofacial demethylation differential expression embryonic stem cell epigenetic regulation gastrulation genome-wide improved in vivo insight interest member nerve stem cell nervous system development nestin protein neurodevelopment neuroregulation new therapeutic target novel programs promoter relating to nervous system response self-renewal skills stem cell biology stem cell biomarkers stem cell function stem cell proliferation targeted treatment transcriptome sequencing transcriptomics whole genome

项目摘要

ABSTRACT The ten-eleven translocation family (TET1/2/3) of enzymes are epigenetic regulators of gene expression that are highly expressed in neural stem cells (NSCs) and during mammalian nervous system development. TET enzymes are dioxygenases that promote active and passive DNA demethylation by converting 5-methylcytosine (5mC) into 5-hydroxymethycytosine (5hmC) and higher-order oxidized derivatives. In addition to its role as a demethylation intermediate, 5hmC can function as a stable epigenetic mark and is highly enriched and dynamic in the developing nervous system. TET enzymes and 5hmC dysregulation have been implicated in human neurodevelopmental syndromes, intellectual disability, craniofacial abnormalities, and neurodegeneration. These observations suggest a critical role for TET enzymes in the developing nervous system and has led to interest in their roles in the biology of NSCs. However, the functions of TET enzymes in NSCs and neurodevelopment remain poorly understood. Preliminary data from our lab demonstrates that Tet triple- knockout NSCs (T123–/–) derived from embryonic stem cells exhibit severe defects in self-renewal, multipotency, and expression of neurodevelopmental genes. We therefore hypothesize that TET enzymes have essential functions in epigenetic regulation of gene expression programs critical for NSC maintenance and multipotency and in embryonic neurodevelopment. To test this hypothesis, we have derived embryonic forebrain NSC lines containing floxed alleles of Tet1/2/3 and a tamoxifen-inducible Cre recombinase transgene expressed from the constitutive Rosa26 locus (T123F/F; +/R26-CreER) for conditional, combined deletion of all three Tet genes. We have also established a colony of Tet1/2/3 triple-floxed mice expressing a tamoxifen-inducible Cre recombinase transgene under control of the neural-specific Nestin promoter (T123F/F; +/Nestin-CreERT2). Using these models, we will (1) define the role of TET enzymes in the maintenance and multipotency of NSCs , (2) establish the requirement of TETs in embryonic neurodevelopment, and (3) identify TET-mediated epigenetic and transcriptional regulatory mechanisms in NSCs. Findings from these studies will define novel roles played by TET enzymes in NSC biology and neurodevelopment, provide insights into how dysregulation of TETs contribute to human neurodevelopmental disorders, and identify novel targets for therapy. Under the joint mentorship of Drs. Meelad Dawlaty and Jean Hébert, I will successfully execute the proposed research and training plan. This research program will further my knowledge of the epigenetic regulation of neural stem cell biology and facilitate my scientific and professional development by equipping me with the necessary skills to become a physician- scientist.

抽象的酶的 10-11 易位家族 (TET1/2/3) 是基因表达的表观遗传调节因子在神经干细胞 (NSC) 和哺乳动物神经系统发育过程中高度表达。酶是双加氧酶，通过转化 5-甲基胞嘧啶来促进主动和被动 DNA 去甲基化 (5mC) 转化为 5-羟甲基胞嘧啶 (5hmC) 和高阶氧化衍生物。去甲基化中间体，5hmC可以作为稳定的表观遗传标记，并且高度富集和动态 TET 酶和 5hmC 失调与人类神经系统发育有关。神经发育综合征、智力障碍、颅面异常和神经变性。这些观察结果表明 TET 酶在发育中的神经系统中发挥着关键作用，并导致然而，TET 酶在 NSC 中的功能和作用令人感兴趣。我们实验室的初步数据表明，Tet 的神经发育仍然知之甚少。源自胚胎干细胞的敲除型 NSC（T123–/–）在自我更新、多能性、因此，我们发现 TET 酶对于神经发育基因的表达至关重要。对 NSC 维持和多能性至关重要的基因表达程序的表观遗传调控功能为了验证这一假设，我们衍生了胚胎前脑 NSC 系。含有 Tet1/2/3 的 floxed 等位基因和由他莫昔芬诱导的 Cre 重组酶转基因组成型 Rosa26 位点（T123F/F；+/R26-CreER）用于条件性、组合删除所有三个 Tet 基因。还建立了表达他莫昔芬诱导型 Cre 重组酶的 Tet1/2/3 Triple-flox 小鼠群体使用这些模型，在神经特异性巢蛋白启动子（T123F/F；+/巢蛋白-CreERT2）的控制下进行转基因。我们将 (1) 定义 TET 酶在 NSC 的维持和多能性中的作用，(2) 建立 TET 在胚胎神经发育中的需求，以及（3）识别 TET 介导的表观遗传和这些研究的结果将定义 NSC 中的转录调控机制。 NSC 生物学和神经发育中的 TET 酶提供了有关 TET 失调如何影响的见解人类神经发育障碍，并在联合指导下确定新的治疗靶点。 Meelad Dawlaty 博士和 Jean Hébert 博士，我将成功执行拟议的研究和培训计划。研究计划将加深我对神经干细胞生物学表观遗传调控的了解，并促进通过使我具备成为一名医生所需的技能来实现我的科学和专业发展- 科学家。