Neuronal Activity-Dependent Regulation of MeCP2

描述（由申请人提供）：概要/摘要：MeCP 2（一种甲基-CpG结合蛋白，作为基因表达的调节剂）中的突变是Rett综合征（RTT）的主要原因，Rett综合征是一种X连锁进行性自闭症谱系障碍，是女孩和妇女严重认知障碍的最常见原因之一。虽然MeCP 2在神经元中的选择性失活已被认为足以在小鼠中赋予Rett样表型，但有丝分裂后神经元中MeCP 2功能丧失导致RTT表型的具体机制仍不清楚。我们已经确定丝氨酸421（S421）MeCP 2作为一个网站的神经元活性依赖性磷酸化，诱导选择性地在大脑中响应生理刺激。值得注意的是，我们发现S421磷酸化控制MeCP 2调节培养神经元和切片制备物中树突图案形成、棘形态发生以及Bdnf转录活性依赖性诱导的能力。为了进一步探索这种调节机制在体内神经发育中的作用，我们已经产生了一种敲入小鼠，其中MeCP 2的S421突变为丙氨酸残基（S421 A KI），防止MeCP 2在该位点的磷酸化。有趣的是，尽管体内MeCP 2 S421磷酸化的消除不会导致MeCP 2表达完全丧失时观察到的运动和存活表型，但我们的初步研究揭示了这些S421 A KI小鼠中皮质抑制性突触发育的缺陷，这表明，活动-依赖性磷酸化可能涉及与RTT中观察到的突触和认知缺陷相关的MeCP 2功能的特定子集。为了开始验证这一假说并确定MeCP 2作为神经元活动依赖性基因表达的一般调节剂的功能，我们提出了以下具体目标：（1）研究MeCP 2 S421磷酸化对体内经验依赖性突触发育的贡献;（2）评估MeCP 2 S421磷酸化在活性依赖性神经元基因表达调节中的作用;（3）表征活性依赖性MeCP 2磷酸化的其他位点。我们希望，拟议的实验将提供一个更好地了解MeCP 2功能，深入了解活性依赖性基因表达的机制，并提供新的机会，为发展治疗策略，以减轻RTT病理。 公共卫生相关性：项目叙述：雷特综合征是一种进行性自闭症障碍，是女孩和妇女严重认知障碍的最常见原因之一。为了深入了解这种疾病的潜在分子基础，这项拟议中的研究将寻求探索这种自闭症疾病反映了中枢神经系统基因动态调节缺陷的假设。

DESCRIPTION (provided by applicant): Summary/Abstract: Mutations in MeCP2, a methyl-CpG-binding protein that functions as a regulator of gene expression, are a major cause of Rett Syndrome (RTT), an X-linked progressive autism spectrum disorder that is among the most common causes of profound cognitive impairment in girls and women. While the selective inactivation of MeCP2 in neurons has been suggested to be sufficient to confer a Rett-like phenotype in mice, the specific mechanisms by which the loss of MeCP2 function in postimitotic neurons contributes to RTT phenotypes remain unclear. We have identified serine 421 (S421) on MeCP2 as a site of neuronal activity-dependent phosphorylation that is induced selectively in the brain in response to physiological stimuli. Significantly, we have found that S421 phosphorylation controls the ability of MeCP2 to regulate dendritic patterning, spine morphogenesis, and the activity-dependent induction of Bdnf transcription in both cultured neurons and slice preparations. To further explore the role of this regulatory mechanism in neural development in vivo, we have generated a knock-in mouse in which S421 of MeCP2 is mutated to an alanine residue (S421A KI), preventing the phosphorylation of MeCP2 at this site. Intriguingly, whereas the abrogation of MeCP2 S421 phosphorylation in vivo does not result in the motor and survival phenotypes seen with complete loss of MeCP2 expression, our preliminary studies have revealed a deficit in cortical inhibitory synaptic development in these S421A KI mice, suggesting that activity-dependent phosphorylation may be involved in a specific subset of MeCP2 functions relevant to the synaptic and cognitive defects observed in RTT. To begin to test this hypothesis and determine the extent to which MeCP2 functions as a general regulator of neuronal activity- dependent gene expression, we propose the following specific aims: (1) to investigate the contribution of MeCP2 S421 phosphorylation to experience-dependent synaptic development in vivo; (2) to assess the role of MeCP2 S421 phosphorylation in the regulation of activity-dependent neuronal gene expression; and (3) to characterize additional sites of activity-dependent MeCP2 phosphorylation. It is our hope that the proposed experiments will provide a better understanding of MeCP2 function, give insight into the mechanisms of activity-dependent gene expression, and provide new opportunities for the development of therapeutic strategies to alleviate RTT pathology. PUBLIC HEALTH RELEVANCE: Project Narrative: Rett Syndrome is a progressive autistic disorder that is among the most common causes of profound cognitive impairment in girls and women. In an effort to gain insight into the underlying molecular basis of the disorder, the proposed study will seek to explore the hypothesis that this autistic disorder reflects a defect in the dynamic regulation of genes in the central nervous system.