Molecular Pathogenesis Studies of Rett Syndrome

描述（由申请人提供）：甲基CpG结合蛋白2（MeCP 2）在二十多年前首次纯化，并被鉴定为结合甲基化CpG二核苷酸的转录抑制因子。11年前，我们的实验室发现X连锁MECP 2基因的突变导致Rett综合征（RTT，MIM 312750）。我们现在知道，MECP 2突变（以及野生型基因的重复或三倍）会导致各种神经精神疾病，从新生儿脑病到自闭症，各种认知和运动障碍，以及男性和女性的早发性精神病。MeCP 2似乎也不是一个简单的转录阻遏物。通过这项资助的最后一次更新所支持的工作，我们令人惊讶地发现，MECP 2过表达增加了80%的下丘脑基因的表达水平，而MeCP 2的缺失导致相同基因的表达降低。更令人困惑的是，染色质免疫沉淀（ChIP）芯片和ChIP-seq数据以及基因座特异性ChIP数据显示，MeCP 2在整个基因组中广泛结合，但它特别集中在特定的启动子上。MeCP 2与启动子或非启动子DNA结合如何增加神经元基因表达尚不清楚。我们假设MeCP 2调节染色质结构，但它也有独特的功能，在选择神经元启动子。因此，在我们的第一个目标中，我们将绘制MeCP 2在大脑中的全基因组占有率，并将结果与我们现有的MeCP 2小鼠模型中的表达数据进行比较，以确定MeCP 2启动子占有率与基因表达变化之间是否存在关系。在我们的第二个目标中，我们将测试MeCP 2作为替代连接组蛋白的假设，并确定脑组织中MeCP 2和H1水平之间是否存在必要的平衡，以及阐明这两个因素之间的功能关系。在第三个目标中，我们将研究MeCP 2与染色质重塑蛋白Ezh 1（我们最近发现的MeCP 2的体内伴侣，其时空表达模式与MeCP 2的时空表达模式相似）的相互作用;在目标2和3中，我们将测试这些相互作用如何通过生成新的小鼠模型来介导MeCP 2表型。最后，我们的第四个目标扩展了我们的工作，表明改变基因表达可能会减轻RTT和MeCP 2重复表型。我们将在RTT和MECP 2复制综合征小鼠模型中测试靶向染色质状态的疗法（HDAC和HAT抑制剂）。这四个目标将解决关于MeCP 2在转录中的作用的几个基本问题，将产生对神经元中染色质改变的重要见解，这些改变在没有体内研究的情况下无法接近，并且可能产生有希望的候选疗法。 公共卫生相关性：MECP 2的突变导致Rett综合征以及许多其他神经精神疾病。我们最近的进展已经导致了可能的治疗方法，我们将在我们的良好表征的小鼠模型中进行测试，同时我们还试图回答有关MeCP 2对神经元基因转录影响的基本问题。

DESCRIPTION (provided by applicant): Methyl-CpG-binding protein 2 (MeCP2) was first purified over twenty years ago and identified as a transcriptional repressor that binds to methylated CpG dinucleotides. Eleven years ago our lab discovered that mutations in the X-linked MECP2 gene cause Rett Syndrome (RTT, MIM312750). We now know that MECP2 mutations (as well as duplications or triplications of the wild-type gene) cause a variety of neuropsychiatric disorders, ranging from neonatal encephalopathy to autism, various kinds of cognitive and motor impairments, and early-onset psychosis in males and females. It also appears that MeCP2 is not a straightforward transcriptional repressor. Through work supported by the last renewal of this grant, we made the surprising discovery that MECP2 overexpression increases the expression levels of 80% of the hypothalamic genes it appears to regulate, whereas loss of MeCP2 results in decreased expression of the same genes. More puzzling still, chromatin immunoprecipitation (ChIP)-chip and ChIP-seq data as well as locus-specific ChIP data show that MeCP2 binds widely throughout the genome but that it is especially concentrated at specific promoters. How MeCP2 binding to promoter or non-promoter DNA increases neuronal gene expression is unclear. We hypothesize that MeCP2 modulates chromatin architecture but that it also has unique functions at select neuronal promoters. In our first aim, therefore, we will map the genome-wide occupancy of MeCP2 in the brain and compare the results with our existing expression data in MeCP2 mouse models to determine if there is a relationship between MeCP2 promoter occupancy and changes in gene expression. In our second aim, we will test the hypothesis that MeCP2 serves as an alternative linker histone, and determine whether there is an essential balance between MeCP2 and H1 levels in brain tissue, as well as elucidate the functional relationship between these two factors. In the third aim, we will examine MeCP2 interactions with the chromatin remodeling protein Ezh1 (an in vivo partner of MeCP2 we recently identified, whose spatiotemporal expression pattern parallels that of MeCP2); in both aims 2 and 3 we will test how these interactions might mediate MeCP2 phenotypes through the generation of new mouse models. Finally, our fourth aim extends our work suggesting that altering gene expression might mitigate RTT and MeCP2 duplication phenotypes. We will test therapies targeting chromatin status (HDAC and HAT inhibitors) in mouse models of RTT and MECP2 duplication syndrome. These four aims will settle several fundamental questions about MeCP2's roles in transcription, will yield important insights into chromatin alterations in neurons that cannot be approached without in vivo studies, and could yield promising candidate therapies. PUBLIC HEALTH RELEVANCE: Mutations in MECP2 cause Rett syndrome as well as a host of other neuropsychiatric disorders. Our recent progress has led to possible therapeutic approaches we will test in our well-characterized mouse models, while we also seek to answer fundamental questions about MeCP2's influence on neuronal gene transcription.