Chromatin Mechanisms of Neuronal Maturation

神经元成熟的染色质机制

• 批准号: 9923467
• 负责人: Anne Elizabeth West
• 金额: $ 48.56万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2017
• 资助国家: 美国
• 起止时间: 2017-05-01 至 2022-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9923467
• 关键词: ARHGEF5 gene; Address; Adult; Biological Models; Brain; CRISPR/Cas technology; Cells; Cerebellum; ChIP-seq; Chromatin; Chromatin Structure; Cues; Cytoplasmic Granules; Data; Development; Distal; Enhancers; Enzymes; Event; Foundations; Gene Activation; Gene Expression; Gene Silencing; Genes; Genetic Transcription; Genomic DNA; Genomics; Goals; Histone H3; Histones; Impairment; Intellectual functioning disability; Kinetics; Lysine; Mediating; Modification; Molecular; Mus; Mutation; Neurodevelopmental Disorder; Neuronal Differentiation; Neurons; Phase; Phosphorylation; Play; Population; Process; Proteins; Recruitment Activity; Regulation; Regulator Genes; Regulatory Element; Repression; Role; Sensory; Synapses; Testing; Time; Transcriptional Activation; Transcriptional Regulation; Variant; autism spectrum disorder; cell type; conditional knockout; critical period; demethylation; derepression; epigenetic regulation; experimental study; gene induction; gene product; genomic locus; histone methyltransferase; in vivo; insight; knock-down; mRNA Expression; mouse model; neuronal circuitry; postnatal; programs; promoter; relating to nervous system; selective expression; synaptic function

Neuronal differentiation requires the precise orchestration of gene expression programs. Temporal control of transcription is particularly important in maturing postmitotic neurons of the postnatal brain because these cells need to express gene products that refine neuronal function and circuitry during specific critical periods of brain development. Epigenetic regulation of chromatin structure is known to play a role in establishing cell-type specific programs of gene expression during early stages of cell fate determination, but the chromatin mechanisms that regulate gene expression during terminal phases of differentiation remain to be fully understood. The goal of this proposal is to identify chromatin mechanisms that coordinate the induction of genes in maturing neurons of the CNS. To discover these chromatin mechanisms of neuronal differentiation, and to determine how these mechanisms are regulated across multiple stages of neuronal maturation, we have characterized chromatin accessibility, enhancer activation, and gene expression in differentiating cerebellar granule neurons (CGNs) of the postnatal mouse in vivo. We have observed that thousands of regulatory elements show chromatin accessibility changes as CGNs differentiate, and we have verified that many of these differentially accessible regions function as developmental stage-specific enhancers of neuronal genes. Furthermore our data identify a specific group of the genes selectively expressed in mature CGNs of the cerebellum that are associated with repressive histone H3 lysine 27 trimethylation (H3K27me3) at early postmitotic stages of CGN differentiation yet lose this mark as CGNs mature. Consistent with a role for active histone demethylation in the induction of these genes, we find that at least a subset of these genes show increased H3K27me3 and reduced mRNA expression in CGNs lacking the H3K27 demethylase Kdm6b. The overarching hypothesis of this proposal is that the molecular regulators of H3K27me3 dynamics in postmitotic neurons play a key role in timing neuronal maturation. To address this hypothesis we propose the following Specific Aims: 1) To characterize the dynamic demethylation of H3K27 in postmitotic neurons maturing in vivo, 2) To determine the role of the H3K27 demethylase Kdm6b in neuronal maturation, and 3) To identify enhancer mechanisms of transcriptional derepression in maturing neurons. Together these studies will identify the molecular mechanisms that mediate the gene-specific loss of H3K27me3 in maturing CGNs, and they will directly test the importance of the developmental derepression of H3K27me3-silenced genes for functional neuronal maturation.

神经元分化需要基因表达程序的精确编排。时间控制 转录对于出生后大脑的有丝分裂后神经元的成熟特别重要，因为这些细胞 需要表达在大脑特定关键时期改善神经元功能和电路的基因产物 发展。已知染色质结构的表观遗传调控在建立细胞类型中发挥作用 细胞命运决定早期阶段基因表达的特定程序，但染色质 在分化末期调节基因表达的机制仍有待充分研究 明白了。该提案的目标是确定协调诱导的染色质机制 中枢神经系统成熟神经元中的基因。为了发现神经元分化的这些染色质机制， 为了确定这些机制如何在神经元成熟的多个阶段进行调节，我们 具有染色质可及性、增强子激活和分化过程中基因表达的特征 出生后小鼠体内的小脑颗粒神经元（CGN）。我们观察到，数以千计的 随着 CGN 的分化，调控元件显示染色质可及性发生变化，我们已经证实 许多这些不同的可接近区域充当神经元发育阶段特异性增强子 基因。此外，我们的数据确定了在成熟 CGN 中选择性表达的一组特定基因 早期与抑制性组蛋白 H3 赖氨酸 27 三甲基化 (H3K27me3) 相关的小脑 CGN 分化的有丝分裂后阶段随着 CGN 的成熟而失去了这一标记。与主动角色一致 组蛋白去甲基化在这些基因的诱导中，我们发现这些基因的至少一个子集显示 在缺乏 H3K27 去甲基化酶 Kdm6b 的 CGN 中，H3K27me3 增加，mRNA 表达减少。这 该提议的总体假设是 H3K27me3 动力学的分子调节因子 有丝分裂后神经元在神经元成熟定时中起着关键作用。为了解决这个假设，我们提出 具体目标如下： 1) 表征有丝分裂后神经元中 H3K27 的动态去甲基化 体内成熟，2) 确定 H3K27 去甲基酶 Kdm6b 在神经元成熟中的作用，以及 3) 确定成熟神经元转录去抑制的增强机制。这些研究将共同 确定介导成熟 CGN 中 H3K27me3 基因特异性丢失的分子机制，以及 他们将直接测试 H3K27me3 沉默基因的发育去抑制的重要性 功能性神经元成熟。