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How does neuronal contact mediate astrocyte transcriptional maturation?

神经元接触如何介导星形胶质细胞转录成熟？

基本信息

批准号：
10748163
负责人：
Justin Savage
金额：
$ 4.06万
依托单位：
DUKE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-09-01 至 2027-08-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10748163
关键词：
Address Adhesions Astrocytes Bar Codes Binding Sites Brain CRISPR-mediated transcriptional activation Cell Adhesion Molecules Cell Nucleus Cellular biology ChIP-seq Chromatin Chromatin Remodeling Factor Coculture Techniques Complex DNA Data Development Doctor of Philosophy EP300 gene Enzymes Epigenetic Process Fibroblasts Flow Cytometry Follow-Up Studies Future Gene Expression Gene Expression Regulation Genes Genetic Transcription Genome Genus Mentha Histones Homeostasis Immunoprecipitation Impairment Infiltration Knock-out Knockout Mice Knowledge Life Link LoxP-flanked allele MLL gene Measures Mediating Methanol Methods Methylation Methyltransferase Modification Molecular Morphogenesis Morphology Mus Nature Neurodevelopmental Disorder Neuroglia Neurons Neuropil Rattus Signal Transduction Synapses Techniques Testing Training Wild Type Mouse Work autism spectrum disorder cell type chromatin immunoprecipitation chromatin modification epigenetic regulation experience experimental study genetic manipulation genomic locus histone methyltransferase histone modification indexing member nerve stem cell neurogenesis neuroligin 2 neuronal circuitry novel overexpression pharmacologic postnatal prevent synaptic function synaptogenesis transcriptome transcriptome sequencing

项目摘要

ABSTRACT Developing a healthy functioning brain depends on the coordination of neuronal circuit formation by glial cells called astrocytes. Previous work has shown that astrocytes, born from the same neural stem cells after the end of neurogenesis, require contact with neurons to mature morphologically, functionally, and transcriptionally. Astrocytic expression of the cell adhesion molecule neuroligin 2 (Nlgn2) is necessary for neuronal contact- dependent astrocyte morphogenesis and synaptogenesis. These findings led us to investigate how neuronal contact mediates astrocyte morphogenesis and synaptogenic functions. To address this question, we performed multiplexed indexed T7 chromatin immunoprecipitation (MintChIP) sequencing to measure histone modifications across the astrocyte genome. These experiments found that the chromatin modification landscape changes significantly over postnatal astrocyte development. Interestingly, when we measured histone modifications in astrocytes from Nlgn2 KO mice, we found a substantial decrease in the H3K4me1 and H3K4me3 modifications compared to wildtype astrocytes. Based on these preliminary findings, this proposal will test the hypothesis that epigenetic histone modifications are the mechanistic link between neuronal contact and astrocyte transcriptional maturation. Specifically, aim 1 will test how histone modifications and transcriptional maturation change when astrocytes are cultured with or without neurons and whether inhibiting histone-modifying enzymes is sufficient to prevent astrocyte transcriptional maturation. Aim 2 will investigate the molecular mechanisms of Nlgn2-dependent astrocyte transcriptional maturation. In particular, we will test the hypothesis that the chromatin remodeler Chd8 is required for Nlgn2-dependent H3K4 methylation and that preventing this H3K4 methylation through astrocyte-specific Chd8 knockout will prevent astrocyte transcriptional maturation. The successful completion of these aims will determine the extent to which histone modifications regulate astrocyte maturation and further our understanding of the molecular mechanism responsible for Nlgn2-induced gene expression changes in astrocytes. These findings will also be highly relevant to future work in neurodevelopmental disorders as astrocyte-regulated synaptogenesis, and epigenetic regulation of gene expression are emerging as prominent aspects of conditions like autism spectrum disorders. Finally, this project will provide an excellent training experience for Mr. Justin Savage through its intersection of epigenetics and glial cell biology.

摘要发展健康的大脑功能有赖于神经胶质细胞对神经元回路形成的协调叫做星形胶质细胞。以前的工作表明，星形胶质细胞出生后，结束于相同的神经干细胞对于神经发生，需要与神经元接触才能在形态、功能和转录上成熟。星形细胞表达细胞黏附分子神经连接蛋白2(Nlgn2)是神经元接触所必需的。依赖星形胶质细胞的形态发生和突触发生。这些发现引导我们研究神经元是如何接触调节星形胶质细胞的形态发生和突触发生功能。为了解决这个问题，我们执行了多重索引T7染色质免疫沉淀(MintChip)测序法测定组蛋白修饰整个星形细胞基因组。这些实验发现，染色质修饰的景观发生了变化显著超过出生后星形胶质细胞的发育。有趣的是，当我们在 Nlgn2KO小鼠星形胶质细胞，我们发现H3K4me1和H3K4me3修饰显著减少与野生型星形胶质细胞相比。基于这些初步发现，这项提议将检验这一假设表观遗传组蛋白修饰是神经元接触和星形胶质细胞之间的机制联系转录成熟。具体地说，目标1将测试组蛋白修饰和转录成熟星形胶质细胞加或不加神经元培养时的变化以及是否抑制组蛋白修饰酶足以阻止星形胶质细胞转录成熟。Aim 2将研究其分子机制。 Nlgn2依赖的星形胶质细胞转录成熟。特别是，我们将测试染色质的假设依赖于Nlgn2的H3K4甲基化需要重构剂CHD8，并防止这种H3K4甲基化通过星形胶质细胞特异性的CHD8基因敲除将阻止星形胶质细胞转录成熟。成功者这些目标的完成将决定组蛋白修饰调控星形胶质细胞成熟的程度。进一步了解了Nlgn2诱导基因表达的分子机制星形胶质细胞的变化。这些发现也将与神经发育障碍的未来工作高度相关。作为星形胶质细胞调节的突触发生和基因表达的表观遗传调节正日益突出自闭症谱系障碍等疾病的各个方面。最后，这个项目将提供一个很好的培训贾斯汀·萨维奇先生通过其交叉的表观遗传学和神经胶质细胞生物学的经验。