Establishment of Neuron-Specific CpG Methylation Patterns During Development in N

描述（由申请人提供）：DNA甲基化与神经元功能和精神障碍有关，但这些研究仅限于少数候选基因。了解DNA甲基化在神经元发育和疾病中的作用需要了解甲基化胞嘧啶在整个基因组和特定神经元原型中的分布，其功能可以直接与正常和病理行为联系起来。在这里，我们建议i）采用全基因组代表性DNA甲基化分析来表征mCpG在海马和纹状体中的谷氨酸能和GABA能投射神经元中的分布，以及在神经元前体和小鼠胚胎干（ES）细胞中的分布，以跟踪发育过程中的甲基化。本研究选择的神经元参与重要功能，包括空间学习/记忆（背侧海马），情绪行为（腹侧海马），运动活动（背侧纹状体）和药物滥用和社会行为（腹侧纹状体/神经核）。由于所有这些行为都受到母体环境的影响，我们还建议ii）确定不利的产前/产后早期母体环境对动物神经元DNA甲基化的影响，这些影响敏感和有弹性。我们先前建立了两种动物模型，其基于母体突变导致遗传正常后代的永久性行为异常，包括异常恐惧、应激反应、整体活动和社会互动。我们期望我们的研究将鉴定DNA甲基化特征，其特异性a）针对神经元原型，B）针对其发育阶段（神经元前体、年轻和成熟神经元），c）针对其背腹位置和d）针对不良母体效应。在这个提议中使用的高分辨率甲基化测定将有助于不仅确定甲基化的总体模式，而且还产生由甲基化标记的基因的组装。这些基因组形成了功能网络和途径，对于理解DNA甲基化在神经元发育和精神疾病中的作用至关重要。 公共卫生相关性：DNA的二级修饰，如胞嘧啶甲基化，可以具有与一级DNA序列中的突变和多态性变异所引起的生物学效应相似的生物学效应。DNA甲基化与精神症状的发展有关，特别是与不良母体环境有关的那些，但这些研究仅限于少数个别候选基因，并使用含有各种神经元和神经胶质细胞的组织进行。在这里，我们提出了全基因组甲基化研究与同质群体的神经元在正常和扰动环境中的发展。

DESCRIPTION (provided by applicant): DNA methylation has been implicated in neuronal functions and mental disorders but these studies were limited to a few candidate genes. Understanding the role of DNA methylation in neuronal development and disease requires knowledge of the distribution of methylated cytosines in the entire genome and in specific neuronal prototypes whose function can be directly linked to normal and pathological behavior. Here we propose i) to employ genome-wide representational DNA methylation analyses to characterize the distribution of mCpGs in glutamatergic and GABAergic projection neurons in the hippocampus and striatum, respectively, as well as in neuronal precursors and mouse embryonic stem (ES) cells to follow methylation during development. The neurons selected for this study are involved in important functions including spatial learning/memory (dorsal hippocampus), emotional behavior (ventral hippocampus), locomotor activity (dorsal striatum) and drug abuse and social behavior (ventral striatum/nucleus accumbens). Since all of these behaviors are influenced by the maternal environment, we also propose ii) to determine the effect of adverse pre/early postnatal maternal environment on neuronal DNA methylation in animals sensitive and resilient to these effects. We have previously established two animal models which are based on maternal mutations leading to permanent behavioral abnormalities in genetically normal offspring, including abnormal fear, stress responsiveness, overall activity and social interaction. We expect that our studies will identify DNA methylation signatures specific a) for neuronal prototypes, b) for their developmental stages (neuronal precursor, young and mature neurons), c) for their dorso-ventral position and d) for adverse maternal effects. The high resolution methylation assay used in this proposal will help to determine not only the overall pattern of methylation but also to generate assemblies of genes marked by methylation. These groups of genes form functional networks and pathways that are essential to understand the role of DNA methylation in neuronal development and psychiatric disease. PUBLIC HEALTH RELEVANCE: Secondary modifications of DNA, such as methylation at cytosines, can have a biological effect similar to those caused by mutations and polymorphic variations in the primary DNA sequence. DNA methylation has been implicated in the development of psychiatric symptoms, especially in those linked to adverse maternal environment, but these studies were limited to a few individual candidate genes and performed with tissue containing various neurons and glia cells. Here we propose whole-genome methylation studies with homogenous populations of neurons during development in normal and perturbed environment.