Molecular Pathogenesis Studies of Rett Syndrome

描述(申请人提供)：甲基CpG结合蛋白2(MeCP2)在20多年前首次被纯化，并被鉴定为一种转录抑制因子，与甲基化的CpG二核苷酸结合。11年前，我们的实验室发现，X连锁MECP2基因的突变会导致Rett综合征(RTT，MIM312750)。我们现在知道MECP2突变(以及野生型基因的复制或三重)会导致各种神经精神障碍，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。似乎MeCP2也不是一个直接的转录抑制因子。通过上一次更新这项拨款所支持的工作，我们有了一个惊人的发现，MECP2的过度表达增加了它似乎调控的80%的下丘脑基因的表达水平，而MeCP2的缺失会导致相同基因的表达减少。更令人费解的是，染色质免疫沉淀(CHIP)-CHIP和CHIP-SEQ数据以及特定位点的芯片数据表明，MeCP2广泛结合在整个基因组中，但它特别集中在特定的启动子上。MeCP2与启动子或非启动子DNA的结合如何增加神经元基因的表达尚不清楚。我们假设MeCP2调节染色质结构，但它也在特定的神经元启动子上具有独特的功能。因此，在我们的第一个目标中，我们将绘制MeCP2在大脑中的全基因组占有率，并将结果与我们在MeCP2小鼠模型中现有的表达数据进行比较，以确定MeCP2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，我们将检验MeCP2作为另一种连接物组蛋白的假设，并确定在脑组织中MeCP2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MeCP2与染色质重塑蛋白Ezh1(我们最近发现的MeCP2的体内合作伙伴，其时空表达模式与MeCP2平行)的相互作用；在两个目标2和3中，我们将测试这些相互作用如何通过产生新的小鼠模型来调节MeCP2的表型。最后，我们的第四个目标扩展了我们的工作，即改变基因表达可能会减少RTT和MeCP2的复制表型。我们将在RTT和MECP2重复综合征小鼠模型中测试针对染色质状态(HDAC和HAT抑制剂)的治疗方法。这四个目标将解决关于MeCP2的S在转录中的角色的几个基本问题，将对神经元染色质的变化产生重要的见解，而不进行体内研究是无法接近的，并可能产生有前景的候选疗法。 公共卫生相关性：MECP2基因突变会导致Rett综合征以及其他一系列神经精神疾病。我们最近的进展导致了可能的治疗方法，我们将在我们特征良好的小鼠模型中测试，同时我们也试图回答关于MeCP2的S对神经元基因转录的影响的基本问题。

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.