MOLECULAR PATHOGENESIS STUDIES OF RETT SYNDROME

项目摘要 Rett综合征（RTT）是一种出生后发病的严重神经发育障碍。由功能丧失引起 X连锁基因甲基CpG结合蛋白2（MECP 2）突变， 通常在生命的前12-18个月，达到预期的里程碑，但随后失去了他们获得的 在一段时间的技能，而不是发展刻板印象，共济失调，失用症，癫痫发作，和一系列其他 症状主要减少MeCP 2蛋白产生量的轻度突变导致轻度 症状需要更长的时间才能显现。RTT的各种小鼠模型（雄性Mecp 2无效，雌性 杂合子，携带特定致病突变的小鼠）忠实地再现了这一自然史 （正常健康、退化和恶化）。引人注目的是，在成年期删除该基因， 表型，而成年裸小鼠中Mecp 2的表达拯救了它，表明MeCP 2具有一些作为- 尚未明确的功能，这对神经元的维持至关重要。虽然基因疗法似乎是最好的 希望，MeCP 2在女性中的镶嵌性质（约50%的神经元将具有野生型MeCP 2）提出了一个困难的问题。 这种方法的挑战，因为太多的MeCP 2也会引起疾病（MECP 2复制综合征[MDS]， 这是每一个位一样毁灭性的）。因此，在我们寻找可行的治疗方案时，我们遵循两个原则： 原则：早期正常发育期提供了一个干预和推迟的机会之窗， 发作，以及MeCp 2丢失和症状出现（或在上下文中症状逆转）之间的滞后 这意味着有一系列的分子事件，如果我们能绘制出它们， 以确定可作为治疗靶点和治疗反应生物标志物的关键基因或途径。 先前发现穹窿脑深部刺激改善RTT小鼠的学习和记忆， 他假设，运动训练在前驱期也可能刺激神经回路， 延缓症状发作事实证明，情况确实如此。我们现在试图追踪 以及在MeCP 2耗尽后但在症状发作前发生的细胞变化，以定义 RTT发病机制和生物标志物的反应，因为我们刺激大脑通过强化训练或提高 亚晶突变背景下的MeCP 2水平。在我们的第一个目标，我们将确定转录 在对强化训练做出反应的任务特异性神经元中发生的变化， 电生理学和形态学。在目标2中，我们将追踪分子、表观遗传和细胞的级联反应， 急性MeCP 2消耗后几周的变化，最终导致神经系统疾病， 功能障碍在目标3中，我们将上调两种不同的RTT小鼠模型中的MeCP 2水平，每种模型都携带 降低MeCP 2水平的突变，以确定改善的程度。

Project Summary Rett syndrome (RTT) is a severe neurodevelopmental disorder with postnatal onset. Caused by loss-of-function mutations in the X-linked gene methyl CpG binding protein 2 (MECP2), girls with classic RTT develop normally for the first ~12-18 months of life, reaching the expected milestones, but then lose their acquired skills over a period of weeks and instead develop stereotypies, ataxia, apraxia, seizures, and a host of other symptoms. Milder mutations that primarily reduce the amount of MeCP2 protein produced cause milder symptoms that take longer to manifest. Various mouse models of RTT (male Mecp2 nulls, female heterozygotes, mice bearing specific disease-causing mutations) faithfully reproduce this natural history (normal health, regression, and deterioration). Strikingly, deletion of the gene in adulthood reproduces the full phenotype, whereas expression of Mecp2 in adult null mice rescues it, demonstrating that MeCP2 has some as- yet-unspecified function that is critical for neuronal maintenance. Although gene therapy would seem the best hope, the mosaic nature of MeCP2 in females (~50% of neurons will have wild-type MeCP2) presents a difficult challenge for this approach, as too much MeCP2 also causes disease (MECP2 duplication syndrome [MDS], which is every bit as devastating). Therefore, in our search for viable therapeutic options, we are guided by two principles: the period of early normal development provides a window of opportunity to intervene and delay onset, and the lag between loss of MeCp2 and appearance of symptoms (or reversal of symptoms, in the context of rescuing MDS) means there is a cascade of molecular events that, if we could but map them, should allow us to identify key genes or pathways that could serve as therapeutic targets and biomarkers of treatment response. Having previously found that forniceal deep brain stimulation improves learning and memory in RTT mice, we hypothesized that motor training during the presymptomatic period might also stimulate the neural circuits and delay symptom onset. Indeed, this proved to be the case. We now seek to trace the cascade of molecular and cellular changes that occur after MeCP2 depletion but before the onset of symptoms to define initiators of RTT pathogenesis and biomarkers of response as we stimulate the brain through intensive training or raise MeCP2 levels in the context of hypomorphic mutations. In our first aim, we will identify the transcriptional changes that take place in task-specific neurons that respond to intensive training with improved function, electrophysiology, and morphology. In aim 2, we will trace the cascade of molecular, epigenetic, and cellular changes that follow for several weeks after acute MeCP2 depletion, and that eventually lead to neurological dysfunction. In aim 3, we will upregulate MeCP2 levels in two distinct RTT mouse models, each carrying mutations that reduce MeCP2 levels, to determine the extent of improvement.