Transcriptome analysis of glia responding to injury

神经胶质细胞对损伤反应的转录组分析

• 批准号: 8565632
• 负责人: Mary Allison Logan
• 金额: $ 23.1万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-06-01 至 2015-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8565632
• 关键词: 4-thiouracil; Acute; Adult; Alzheimer' s Disease; Amyotrophic Lateral Sclerosis; Animals; Astrocytes; Axon; Axotomy; Biological Assay; Biological Process; Brain; Brain Injuries; Cells; Cellular biology; Communities; Development; Disease; Drosophila genus; Drosophila melanogaster; Employee Strikes; Event; Gene Expression; Gene Expression Profile; Genes; Genetic; Grant; Health; Human; Immune; Immune response; Injury; Invertebrates; Label; Link; Mammals; Methods; Mission; Molecular; Molecular Profiling; Morphology; Multiple Sclerosis; Nerve Degeneration; Nervous System Physiology; Neurodegenerative Disorders; Neuroglia; Neurologic; Neurons; Organism; Play; Process; Protein Isoforms; Public Health; RNA; RNA Interference; RNA Splicing; Receptor Signaling; Role; Signal Pathway; Signal Transduction; Site; Stress; Surface; Synapses; Transgenic Organisms; Trauma; United States National Institutes of Health; Up-Regulation; Validation; Vertebrates; Work; cell type; cognitive function; deep sequencing; falls; fitness; fly; genetic profiling; in vivo; injured; innate immune function; innovation; insight; interest; migration; nerve injury; nervous system disorder; novel; programs; public health relevance; research study; response; response to injury; therapy development; tool; transcriptome sequencing

DESCRIPTION (provided by applicant): Glia are the most abundant cells in the human brain and they play key roles in CNS function and health. Glial cells regulate synaptic signaling, ensheath axonal projections and, importantly, protect the brain by serving as the first line of defense against neuronal damage. The adult brain contains a striking array of diverse glial subtypes, but little is known about the unique genetic profiles of distinct classes of glia that alow them to carry out their important and varied functions. Moreover, determining how the transcriptional profile of glial cells are altered in response to neural injury has presented a unique set of challenges, since the process of isolating glia from the brain for transcriptional analysis is, in and of itself, highly stressful to the cells. Recent work has shown that the adult Drosophila melanogaster brain contains a variety of glial subtypes that are strikingly similar to those described in vertebrates. In addition, acute neural injury induces glial immune responses in flies that are highly reminiscent of those triggered in mammalian glia, including upregulation of essential glial immune genes. This project will take advantage of these evolutionarily conserved features of glia and integrate cutting-edge advances in the fields of in vivo RNA labeling and high throughput deep sequencing to generate a comprehensive transcriptome of Drosophila glial cells in the intact adult brain before and after injury. We will use novel genetic drivers that are expressed in discrete glial subtypes in the adult fly brain to genetically "label" RNA in each class of glia in vivo and then biochemically isolate the labeled RNA to sequence glial subtype transcriptomes by RNA-seq. Using a well-established axotomy assay, we will perform these experiments in uninjured and injured flies to compare the transcriptional profiles of glia before and after acute axon injury. Finally, we will validate the expression of glial genes identified by RNA-seq and begin to characterize the functional role of the newly discovered immune genes that are acutely regulated in glia responding to axotomy. This work (a) will provide critical mechanistic insight into the function of diverse glial subtypes in the adult brain (b) offers a unique opportunity to investigate how gene expression is altered in glia responding to neurodegeneration in the intact CNS and (c) will generate a valuable genetic toolkit for the scientific community to investigate many unexplored aspects of glial cell biology.

描述（由申请人提供）：胶质细胞是人类大脑中最丰富的细胞，它们在CNS功能和健康中发挥关键作用。神经胶质细胞调节突触信号传导，包裹轴突投射，并且重要的是，通过充当针对神经元损伤的第一道防线来保护大脑。成年人的大脑中含有一系列引人注目的不同胶质细胞亚型，但人们对不同类别的胶质细胞的独特遗传特征知之甚少，这些胶质细胞使它们能够执行其重要而多样的功能。此外，确定神经胶质细胞的转录谱如何响应于神经损伤而改变已经提出了一组独特的挑战，因为从大脑中分离神经胶质用于转录分析的过程本身对细胞具有高度压力。最近的研究表明，成年黑腹果蝇的大脑中含有各种胶质细胞亚型，这些亚型与脊椎动物中描述的胶质细胞亚型惊人地相似。此外，急性神经损伤诱导果蝇的神经胶质免疫反应，这与哺乳动物神经胶质中引发的免疫反应非常相似，包括上调必要的神经胶质免疫基因。该项目将利用神经胶质细胞的这些进化上保守的特征，并整合体内RNA标记和高通量深度测序领域的前沿进展，以在损伤前后的完整成年大脑中生成果蝇神经胶质细胞的全面转录组。我们将使用新的基因 在成年果蝇大脑中以离散的神经胶质亚型表达的驱动程序， RNA在体内的每一类胶质细胞，然后生化分离标记的RNA序列胶质细胞亚型转录组通过RNA-seq。使用一个完善的轴突切开试验，我们将在未受伤和受伤的苍蝇进行这些实验，比较神经胶质细胞的转录谱之前和之后的急性轴突损伤。最后，我们将验证胶质基因的表达 通过RNA-seq鉴定，并开始表征新发现的免疫基因的功能作用，这些免疫基因在神经胶质细胞对轴突切断术的反应中受到急性调节。这项工作（a）将提供关键的机制洞察不同的神经胶质细胞亚型的功能在成年人的大脑（B）提供了一个独特的机会，研究如何改变基因表达的神经胶质细胞在完整的中枢神经系统的神经变性和（c）将产生一个有价值的遗传工具包，为科学界研究神经胶质细胞生物学的许多未开发的方面。