DNA Hypomethylation and Cortical Neuronal Degeneration

描述(申请人提供)：在哺乳动物中，DNA胞嘧啶甲基化是调节许多细胞事件的主要表观遗传因素之一，包括发育基因表达和基因组印记。DNA甲基化机制的改变与几种智力低下障碍有关，包括Rett、ICF、Fragile-X和ATRX综合征，这表明甲基化对神经元的发育和功能很重要。为了研究DNA甲基化在中枢神经系统(CNS)中的作用，我们最近应用cre/loxP系统，专门删除了中枢神经系统中的维持DNA甲基转移酶Dnmt1基因。通过将Emx1-cre转基因与Dnmt1条件等位基因(Dnmt2lox)杂交，我们获得了局限于大脑皮层和海马区的Dnmt1缺陷条件性基因敲除小鼠。Emx1-cre介导的Dnmtl基因缺失在胚胎第9-10天的大脑皮质前体细胞启动，导致胚胎和出生后皮质投射神经元的DNA低甲基化。突变小鼠在成年后仍可存活，但表现出明显的行为缺陷，如多动和尾部悬吊时后肢紧扣。形态学研究表明，Emx1-cre；Dnmtl突变小鼠表现出大量的皮质体积丢失，因此成为研究DNA低甲基化对皮质变性影响的有价值的动物模型。在这项建议中，我们计划首先检查皮质神经细胞死亡的时间进程，并确定突变小鼠的严重组织学和行为缺陷。进一步的实验旨在确定DNA低甲基化对前体细胞的增殖和分化的影响，以及Dnmtl-/-神经元的树突分枝。最后，我们计划确定DNA低甲基化导致突变皮层神经细胞死亡的机制。众所周知，DNA甲基化水平随着年龄的增长而降低，在阿尔茨海默病等与年龄相关的神经退行性疾病中也是如此。了解低甲基化皮质神经元细胞死亡的机制可能有助于我们制定预防低甲基化诱导的神经元变性的治疗策略。

DESCRIPTION (provided by applicant): In mammals DNA cytosine methylation is one of the major epigenetic factors that regulate many cellular events including developmental gene expression and genomic imprinting. Alternations in DNA methylation machinery have been linked to several mental retardation disorders, including Rett, ICF, Fragile-X, and ATRX syndromes, suggesting that methylation is important for neuronal development and function. To investigate the function of DNA methylation in the central nervous system (CNS), we have recently applied the cre/loxP system to delete the maintenance DNA methyltransferase Dnmtl gene exclusively in the CNS. By crossing the Emx1-cre transgene with the Dnmtl conditional allele (Dnmt2lox), we have obtained conditional knockout mice with Dnmtl deficiency restricted to the cortex and hippocampus. Emx1-cre mediated Dnmtl gene deletion is initiated in pallial cortical precursor cells at embryonic day (E) 9-10, resulting in DNA hypomethylation in embryonic and postnatal cortical projection neurons. Mutant mice are viable in adulthood but exhibit obvious behavioral defects such as hyperactivity and hind limb clasping upon tail suspension. Morphological studies indicate that Emx1-cre; Dnmtl mutant mice exhibit massive loss of cortical volume, thus become a valuable animal model for studying the effect of DNA hypomethylation on cortical degeneration. In this proposal, we plan to first examine the time course of cortical neuronal cell death and determine the gross histological and behavioral defects in the mutant mice. Further experiments are designed to determine the effect of DNA hypomethylation on the proliferation and differentiation of precursor cells, as well as dendritic arborization of Dnmtl-/- neurons. Finally, we plan to determine the mechanism by which DNA hypomethylation induces neuronal cell death in the mutant cortex. It is known that levels of DNA methylation decrease with aging and in age-related neurodegenerative disorders such as Alzheimer's disease. Understanding the mechanism of cell death in hypomethylated cortical neurons may help us develop therapeutic strategies to prevent hypomethylation-induced neuronal degeneration.