Neuronal Epigenomic Changes in Neurodevelopment and Disease

描述(由申请人提供)：早期的经验和环境条件，特别是那些发生在大脑可塑性增强时期的条件，已知会促进生理和行为的长期变化，这些变化独立于DNA代码的变化而起作用。越来越多的证据表明，表观基因组过程在这些表观遗传现象中扮演着重要的角色。特别是，来自各种来源的最新数据表明，DNA甲基化的经验依赖性变化可以通过对神经元基因表达的持续影响对神经功能产生长期影响。然而，这些修饰在体内发生在高度异质性神经组织中的相对较少的细胞中，这一事实限制了现有基因组方法对这些修饰的研究，并极大地复杂化了潜在分子机制的研究。我们提出了一个双管齐下的方法来开始解决这些问题。首先，我们将采用一种简化论的方法来研究经验驱动的DNA甲基化变化，使用一个分离的神经元培养系统，该系统显示活动诱导的DNA甲基化变化，其中大量细胞可以在强劲的刺激下同步激活。此外，为了补充和解决这种简化论方法固有的局限性，我们还开发了一种通用的遗传策略，专门从体内定义的细胞类型中分离染色质，从而能够使用大规模平行测序技术分析特定神经细胞群体中因早期生活经验而诱导的DNA甲基化变化。因此，我们建议：1)使用分离的神经元培养系统来表征神经元活动诱导的DNA甲基化的变化，以及2)在活体中研究长期持续的神经元表观基因组与经验驱动的行为和生理变化的相关性。我们希望所提出的实验将为神经元表观基因组修饰的分析建立新的方法，促进我们对发育中的中枢神经系统中DNA甲基化调控的理解，并最终为这些机制对神经发育、认知行为和疾病的重要性提供新的见解。 公共卫生相关性：众所周知，不良的早期生活事件会影响神经发育和精神障碍的风险，引发生理和行为的长期变化，这些变化不依赖于DNA代码的变化。为了深入了解这些效应的潜在分子基础，这项拟议的研究将探索受调控的DNA甲基化在神经元基因表达持续变化中的作用。

DESCRIPTION (provided by applicant): Early-life experiential and environmental conditions, particularly those occurring during heightened periods of brain plasticity, are known to promote long-term changes in physiology and behavior that act independently of changes in the DNA code. Accumulating evidence suggests an important role for epigenomic processes in these epigenetic phenomena. In particular, recent data from a variety of sources suggest that experience-dependent changes in DNA methylation can have a long-lasting impact on neural function through sustained effects on neuronal gene expression. However, the fact that these modifications occur in vivo in a relatively small number of cells within highly heterogeneous neural tissue limits the study of these modifications with existing genomic approaches and greatly complicates investigation of the underlying molecular mechanisms. We propose a two-pronged approach to begin to address these issues. First, we will pursue a reductionist approach to the study of experience-driven changes in DNA methylation, employing a dissociated neuronal culture system that shows activity-induced changes in DNA methylation and in which a large number of cells can be synchronously activated with robust stimuli. Moreover, to complement and address limitations inherent in this reductionist approach, we have also developed a general genetic strategy to specifically isolate chromatin from defined cell types in vivo, enabling the analysis of DNA methylation changes induced in specific neuronal cell populations in response to early-life experiences using massively parallel sequencing techniques. Thus, we propose: 1) To employ a dissociated neuronal culture system to characterize neuronal activity-induced changes in DNA methylation, and 2) To investigate long-lasting neuronal epigenomic correlates to experience-driven behavioral and physiological changes in vivo. It is our hope that the proposed experiments will establish new approaches for the analysis of neuronal epigenomic modifications, advance our understanding of the regulation of DNA methylation in the developing central nervous system, and ultimately provide new insights into the importance of these mechanisms for neurodevelopment, cognitive behavior, and disease. PUBLIC HEALTH RELEVANCE: Adverse early life events are known to influence risk for neurodevelopmental and psychiatric disorders, triggering long-lasting changes in physiology and behavior that act independent of changes to the DNA code. In an effort to gain insight into the underlying molecular basis of these effects, the proposed study will explore the role of regulated DNA methylation in the persistent alteration of neuronal gene expression.