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。理解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，分别表征海马和纹状体中谷氨酸能和氨基丁酸能投射神经元以及神经元前体和小鼠胚胎干（ES）细胞中mcpg的分布，以跟踪发育过程中的甲基化。本研究选择的神经元涉及空间学习/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/伏隔核）等重要功能。由于所有这些行为都受到母体环境的影响，我们还提出ii)确定不利的产前/产后早期母体环境对这些影响敏感和有弹性的动物神经元DNA甲基化的影响。我们之前已经建立了两种动物模型，它们基于母亲突变导致基因正常的后代出现永久性行为异常，包括异常的恐惧、压力反应、整体活动和社会互动。我们期望我们的研究将确定特定于a)神经元原型的DNA甲基化特征，b)它们的发育阶段（神经元前体，年轻和成熟神经元），c)它们的背-腹侧位置和d)母体不良影响。本提案中使用的高分辨率甲基化测定不仅有助于确定甲基化的总体模式，还有助于生成甲基化标记的基因组装。这些基因组形成的功能网络和通路对于理解DNA甲基化在神经元发育和精神疾病中的作用至关重要。

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.