Regulation of the Reelin Gene

描述(申请人提供)：在胚胎大脑发育过程中，卷起 由Cajal-Retzius(CR)细胞分泌到细胞外基质(ECM)中 用于协调迁移神经元在发育过程中的位置 大脑皮层、海马体和小脑。在成年大鼠的大脑皮层中，Reelin是 从GABA能神经元亚群分泌到细胞外基质中 本构机制。在成年的非人类灵长类动物中，卷轴存在于 ECM，与含有α3的整合素受体在邻近 树枝状刺。这增加了成人卷轴蛋白整合素 相互作用可能通过调节表达而促进突触的可塑性 促进树突棘滋养的细胞骨架蛋白。在……里面 精神分裂症患者的死后脑、树突棘密度 是减少的。此外，在从被诊断为脑部疾病的患者获得的大脑中 精神分裂症和伴有精神病的双相情感障碍，reelin基因和蛋白 水平降低了大约50%。基因组中的胞嘧啶甲基化 DNA影响细胞过程的全景，包括基因转录， 基因组印迹和基因组稳定性。这篇文章中概述的实验 应用程序将对支持我们假设的初步数据进行扩展 人类卷轴蛋白启动子是由DNA甲基化调节的，这是 通过染色质的变化决定转录因子的可及性 结构。我们的目标是获得与负责机制相关的信息 寻找合适的reelin基因表达的时间和空间模式。我们 将定义启动子在调节卷绕中起作用的部分 在原代培养的神经细胞(皮质神经元和胶质细胞以及 小脑颗粒神经元)。使用这些信息，我们将确定 与这些调控区域相互作用的反式作用因子(目标1)。 其次，我们还将重点介绍DNA甲基化在定义 Reelin基因(RELN)在体外分化神经前体细胞中的表达 (目标2)。最后，我们将使用reelin启动子产生转基因动物。 驱动LacZ报告基因的表达并检测这些基因的作用 靶向神经元表达RELN的调控序列(目标1)。我们会 检测人类启动子转基因在不同环境中的甲基化模式 来自这些转基因小鼠的原代培养(目标2)，并将比较 内源性基因甲基化与人类卷轴的关系 转基因。从拟议的实验中获得的信息将提供 制定与收卷机构相关的假设的框架 在精神疾病中，表达可能在功能上受到损害。最终， 对负责调节卷轴表达的事件的理解可能 提供关于基因未来可能如何被操纵的初步线索 作为治疗这些复杂精神疾病的一种潜在方法。

DESCRIPTION (provided by applicant): During embryonic brain development, reelin is secreted by Cajal-Retzius (CR) cells into the extracellular matrix (ECM) and serves to orchestrate the positioning of migrating neurons in the developing cortex, hippocampus and cerebellum. In the cortex of adult rat, reelin is secreted from a subpopulation of GABAergic neurons into the ECM by a constitutive mechanism. In adult non-human primates, reelin, present in the ECM, is contiguous with alpha3-containing integrin receptors in the vicinity of dendritic spines. This raises the possibility that adult reelin-integrin interactions may contribute to synaptic plasticity by modulating the expression of cytoskeletal proteins that facilitate the trophism of dendritic spines. In the post-mortem brains of schizophrenia patients, basal dendritic spine density is decreased. Moreover, in brains obtained from patients diagnosed with schizophrenia and bipolar illness with psychosis, reelin mRNA and protein levels are reduced by approximately 50 percent. Cytosine methylation of genomic DNA influences a panorama of cellular processes, including gene transcription, genomic imprinting, and genome stability. Experiments outlined in this application will expand upon our preliminary data that support our hypothesis that the human reelin promoter is modulated by DNA methylation, which determines transcription factor accessibility through alterations in chromatin structure. Our goal is to obtain information relevant to mechanisms responsible for appropriate temporal and spatial patterns of reelin gene expression. We will define those parts of the promoter operative in modulating reelin expression in primary neuronal cell cultures (cortical neurons and glia and cerebellar granule neurons). Using this information, we will identify trans-acting factors that interact with these regulatory regions (Aim 1). Secondly, we will also focus on the role that DNA methylation plays in defining reelin gene (RELN) expression in neuronal precursors differentiating in vitro (Aim 2). Finally, we will generate transgenic animals using the reelin promoter to drive expression of a lacZ reporter gene and examine the role of these regulatory sequences in targeting neuronal expression of RELN (Aim 1). We will examine the methylation pattern of the human promoter transgene in various primary cultures derived from these transgenic mice (Aim 2) and will compare the patterns of methylation of the endogenous gene with the human reelin transgene. Information obtained from the proposed experiments will provide the framework for formulating hypotheses relevant to mechanisms by which reelin expression may be functionally compromised in psychiatric diseases. Ultimately, an understanding of events responsible for modulating reelin expression may provide preliminary clues as to how the gene might be manipulated in the future as one potential therapeutic approach to these complex mental disorders.