Regulation of the Reelin Gene

描述（由申请人提供）：在胚胎大脑发育过程中，reelin 由 Cajal-Retzius (CR) 细胞分泌到细胞外基质 (ECM) 中， 用于协调发育中迁移神经元的定位 皮质、海马体和小脑。在成年大鼠的皮质中，reelin 是 由 GABA 能神经元亚群分泌到 ECM 中 的构成机制。在成年非人类灵长类动物中，reelin 存在于 ECM，与含 α3 的整联蛋白受体相邻 树突棘。这提出了成人卷轴整合素的可能性 相互作用可能通过调节表达来促进突触可塑性 促进树突棘营养的细胞骨架蛋白。在 精神分裂症患者死后大脑，基底树突棘密度 减少了。此外，在从诊断患有以下疾病的患者身上获取的大脑中 精神分裂症和双相情感障碍伴精神病、reelin mRNA 和蛋白质 水平降低约 50%。基因组胞嘧啶甲基化 DNA 影响整个细胞过程，包括基因转录、 基因组印记和基因组稳定性。本文概述的实验 应用程序将扩展支持我们假设的初步数据 人类 reelin 启动子受 DNA 甲基化调节， 通过染色质的改变决定转录因子的可及性 结构。我们的目标是获取与负责机制相关的信息 了解 reelin 基因表达的适当时间和空间模式。我们 将定义启动子的那些可调节卷轴的部分 在原代神经元细胞培养物（皮质神经元和神经胶质细胞）中表达 小脑颗粒神经元）。使用这些信息，我们将识别 与这些调节区域相互作用的反式作用因子（目标 1）。 其次，我们还将关注 DNA 甲基化在定义中所起的作用。 reelin 基因 (RELN) 在体外分化的神经元前体中的表达 （目标 2）。最后，我们将使用 reelin 启动子生成转基因动物 驱动 lacZ 报告基因的表达并检查这些基因的作用 靶向 RELN 神经元表达的调控序列（目标 1）。我们会 检查各种人类启动子转基因的甲基化模式 来自这些转基因小鼠的原代培养物（目标 2）并将进行比较 人类 reelin 内源基因的甲基化模式 转基因。从拟议的实验中获得的信息将提供 制定与卷绕机制相关的假设的框架 在精神疾病中表达可能会受到功能损害。最终， 了解负责调节 reelin 表达的事件可能 为未来如何操纵该基因提供初步线索 作为这些复杂精神障碍的一种潜在治疗方法。

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.