Investigation of MeCP2 Function in Rett Syndrome

描述(由申请人提供)：编码甲基CpG结合蛋白2(MeCP2)的基因的从头突变是Rett综合征(RTT)的主要原因，RTT是一种X连锁的显性神经发育障碍。MeCP2是一种染色质相关蛋白，长期以来一直被认为是甲基化DNA的非特异性转录抑制因子。然而，最近的数据表明，MeCP2可以与特定的转录调控因子相互作用，改变上下文基因的表达。与RTT患者明显的神经学表型一致，新的证据表明MeCP2活性与神经元功能密切相关。在哺乳动物神经系统中，MeCP2在有丝分裂后神经元中表达最高，新发现的剪接变异体Mecp2e1是主要的转录本。MeCP2基因突变导致的表型和神经病理学上的相似性表明，MeCP2的神经功能在人类和小鼠之间是保守的。这项建议的目的是更好地了解MeCP2在培养的神经细胞细胞核中的调节和活性，并确定突变和丝氨酸磷酸化如何影响其功能。这项工作将使用活细胞成像方法来检测MeCP2在细胞核中的流动性，并探索突变、染色质重塑和磷酸化对染色质结合的影响。我们将具体描述一种新发现的蛋白质剪接变体，它是疾病中的相关异构体。我们还将描述神经生长因子和靶神经在交感神经元MeCP2表达和功能调节中的作用，并确定MeCP2缺乏如何影响交感神经元对生理提示的反应能力。总而言之，这些研究应该回答交感神经元通过异构体特异性表达和翻译后修饰来调节MeCP2功能的基本问题。这些研究将有助于阐明MeCP2蛋白的调控方式，并测试致病突变影响蛋白功能的方式。长期目标是了解RTT患者神经系统功能障碍的发病机制，并确定调节该系统的方法，以潜在地改善功能并开发靶向治疗。

DESCRIPTION (provided by applicant): De novo mutations in the gene encoding methyl CpG binding protein 2 (MeCP2) are the primary cause of Rett Syndrome (RTT), an X-linked dominant neurodevelopmental disorder. Mecp2 is a chromatin-associated protein that has long been considered to be a non-specific transcriptional repressor of methylated DNA. However, recent data indicate that Mecp2 can interact with specific transcriptional regulators to alter contextual gene expression. Consistent with the pronounced neurological phenotype in RTT patients, emerging evidence suggests a close association between Mecp2 activity and neuronal function. In the mammalian nervous system, Mecp2 expression is highest in postmitotic neurons, where a newly identified splice variant, Mecp2e1, is the predominant transcript. Similarities in phenotype and neuropathology caused by mutations in Mecp2 suggest that the neuronal function of Mecp2 is conserved between humans and mice. The goals of this proposal are to better understand the regulation and activity of Mecp2 in the nucleus of cultured neuronal cells and determine how mutation and serine phosphorylation affect its function. The work proposed will use live cell imaging approaches to examine the mobility of Mecp2 in the nucleus and explore the consequences of mutation, chromatin remodeling and phosphorylation on chromatin binding. We will specifically characterize a newly identified splice variant of the protein that is the relevant isoform in the disease. We will also characterize the role of nerve growth factor and target innervation in regulation of Mecp2 expression and function in sympathetic neurons and determine how Mecp2 deficiency affects the ability of a sympathetic neuron to respond to physiologic cues. Together these studies should answer fundamental questions about the ways that sympathetic neurons modulate Mecp2 function, through isoform specific expression and post-translational modification. These studies will help clarify the way that the Mecp2 protein is regulated and test the ways in which disease causing mutations affect the protein's function. The long term goals are to understand the pathogenesis of the dysfunction of the nervous system in patients with RTT and determine ways that the system can be modulated to potentially improve function and develop targeted therapies.