Transcriptional condensates, epigenetic editing and Rett Syndrome

转录凝聚体、表观遗传编辑和雷特综合征

• 批准号: 10462550
• 负责人: RUDOLF JAENISCH
• 金额: $ 89.26万
• 依托单位: WHITEHEAD INSTITUTE FOR BIOMEDICAL RES
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-15 至 2024-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10462550
• 关键词: 3-Dimensional; Adult; Affect; Alleles; Architecture; Binding; Binding Sites; Biochemical Reaction; Biological Assay; Brain; Cell physiology; Cells; Chimeric Proteins; Chromatin; Chromosomes; Clinical Trials; Complex; Cultured Cells; DNA; DNA Binding Domain; DNA Sequence Alteration; DNA-Binding Proteins; Data; Development; Disease; Disease Progression; Disease model; EP300 gene; Epigenetic Process; Euchromatin; Female; Gene Expression; Generations; Genes; Genetic Transcription; Goals; Heterochromatin; Human; In Vitro; Link; Liquid substance; MeCP2 Duplication Syndrome; Mediating; Mental Retardation; Methyl-CpG-Binding Protein 2; Mus; Mutant Strains Mice; Mutation; Neurodevelopmental Disorder; Neurons; Patients; Phase; Phenotype; Physiological; Protein Region; Proteins; Psyche structure; Recombinants; Reporter; Rett Syndrome; Role; Site; Symptoms; Syndrome; Testing; Therapeutic; Therapeutic Intervention; Time; Transcription Coactivator; Transcription Repressor; Transgenes; Transgenic Mice; X Inactivation; aged; autism spectrum disorder; base; behavior influence; candidate identification; cohesin; defined contribution; design; disability; disease-causing mutation; early childhood; embryonic stem cell; gene repression; genetic corepressor; girls; human embryonic stem cell; in vivo; insight; mouse development; mutant; novel strategies; novel therapeutic intervention; novel therapeutics; overexpression; postnatal; promoter; protein function; side effect; therapeutically effective; tool; vector

Abstract Rett syndrome (RTT) is a postnatal progressive neurodevelopmental disorder associated with severe mental disability and autism-like syndromes that manifests in girls during early childhood, and is caused by mutation of the X-linked DNA binding protein MeCP2 (Methyl CpG-binding Protein 2). Mice carrying null alleles of Mecp2 closely mimic symptoms seen in patients and are faithful models of the disease. Importantly, development of RTT-like symptoms can be slowed or even halted in the adult following correction of a mutant Mecp2 allele by transgene-mediated MeCP2 expression. MeCP2 is one of the most abundant proteins in neurons, and most disease-causing mutations cluster in the DNA binding domain (MBD) and in the transcription repression domain (TRD). However, the function of MeCP2 remains enigmatic, with two major hypotheses having been proposed: (i) MeCP2 acts as repressor of transcription or (ii) as an activator of transcription. Clearly, none of these proposed functions can fully explain the complex phenotype of MeCP2 deficiency or overexpression leading to RTT or MECP2 Duplication Syndrome. Based on our preliminary evidence we postulate that MeCP2’s primary function may be to modulate the 3D chromosome architecture through condensate formation. Components of both euchromatin and heterochromatin can form phase-separated condensates, which provide a mechanism to compartmentalize and concentrate biochemical reactions within cells and are produced by liquid-liquid phase separation driven by intrinsically disordered regions (IDRs) of proteins. MeCP2 protein contains a large IDR and we have obtained preliminary evidence that MeCP2 is involved in phase- separated heterochromatin condensates. Thus, beyond MeCP2’s role as a repressor or activator of gene expression, the protein may have a much wider and more complex role in the cell physiology and disease. In this project we will define the contribution of MeCP2 to heterochromatic and euchromatic condensates in normal and mutant neurons and analyze the effect of RTT causing mutations on LLPS. Our goal is to gain insights into the function of MeCP2 as the basis for designing novel therapeutic approaches. Potential new therapies based on this hypothesis will take time to develop into applications. To explore a more immediate approach we will use epigenetic editing as a therapeutic tool to activate the inactive wt MECP2 allele located on the inactive X chromosome. Most importantly, epigenetic editing will restore MeCP2 expression to exactly wild type levels and thus avoid toxic consequences of MeCP2 overexpression. In contrast, other strategies such as using vector-mediated MeCP2 transduction will invariably produce cells that overexpress MeCP2 and thus will result in serious side effects as seen in patients with MECP2 duplication syndrome.

抽象的 RETT综合征（RTT）是一种与严重精神有关的产后进行性神经发育障碍 残疾和自闭症综合症在童年时期出现女孩，是由突变引起的 X连锁的DNA结合蛋白MECP2（甲基CpG结合蛋白2）。携带MECP2无效等位基因的小鼠 在患者中看到的紧密模仿符号，并且是该疾病的忠实模型。重要的是，发展 通过校正突变的MECP2等位基因后，成人可以放慢甚至停止RTT样症状 转基因介导的MECP2表达。 MECP2是神经元中最丰富的蛋白质之一，最引起疾病的突变聚类在该神经元中 DNA结合结构域（MBD）和转录表达结构域（TRD）。但是， MECP2仍然神秘，提出了两个主要假设：（i）MECP2充当复制品 转录或（ii）作为转录激活剂。显然，这些提议的功能都无法完全解释 MECP2缺乏或过表达的复杂表型导致RTT或MECP2重复 综合征。根据我们的初步证据，我们认为MECP2的主要功能可能是 通过冷凝物形成调节3D染色体体系结构。 白染色质和异染色质的成分都可以形成相位分离的冷凝物，该冷凝水 提供一种机制，使细胞内的生化反应分隔和浓缩生化反应 由蛋白质本质上无序区域（IDR）驱动的液态液相分离产生。 MECP2 蛋白质包含一个大的IDR，我们获得了MECP2参与相的初步证据 分离的异染色质冷凝物。除了MECP2作为基因复制因子或激活因子的角色之外 表达，蛋白质在细胞生理和疾病中可能具有更广泛，更复杂的作用。 在这个项目中，我们将定义MECP2对异质和构体冷凝物的贡献 正常和突变神经元，分析引起突变对LLP的RTT的影响。我们的目标是获得 洞悉MECP2作为设计新型治疗方法的基础的功能。潜在的新 基于此假设的疗法将需要一些时间才能开发到应用程序中。探索更直接的 方法我们将使用表观遗传编辑作为一种治疗工具，以激活位置的无效WT MECP2等位基因 在非活动X染色体上。最重要的是，表观遗传编辑将使MECP2表达精确地恢复 野生型水平，从而避免MECP2过表达的有毒后果。相反，其他策略 例如使用矢量介导的MECP2转移将总是产生过表达MECP2和的细胞 因此，在MECP2重复综合征患者中会产生严重的副作用。