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 雷特综合征（RTT）是一种产后进行性神经发育障碍，与严重的精神障碍相关。 残疾和自闭症样综合症，在儿童早期出现，由基因突变引起 X连锁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 与患者的症状非常相似，是该疾病的忠实模型。重要的是，发展 通过纠正突变 Mecp2 等位基因后，成人的 RTT 样症状可以减缓甚至停止 转基因介导的 MeCP2 表达。 MeCP2 是神经元中最丰富的蛋白质之一，大多数致病突变集中在 DNA 结合域 (MBD) 和转录抑制域 (TRD)。然而，函数 MeCP2 仍然是个谜，人们提出了两个主要假设：(i) MeCP2 作为 转录或（ii）作为转录激活剂。显然，这些提出的函数都不能完全解释 MeCP2 缺乏或过度表达的复杂表型导致 RTT 或 MECP2 重复 综合症。根据我们的初步证据，我们假设 MeCP2 的主要功能可能是 通过凝结物形成调节 3D 染色体结构。 常染色质和异染色质的成分可以形成相分离的冷凝物， 提供了一种划分和集中细胞内生化反应的机制， 由蛋白质本质无序区域 (IDR) 驱动的液-液相分离产生。甲基CP2 蛋白质含有一个大的IDR，我们已经获得了MeCP2参与相-的初步证据。 分离异染色质浓缩物。因此，除了 MeCP2 作为基因抑制子或激活子的作用之外 表达后，该蛋白可能在细胞生理学和疾病中发挥更广泛和更复杂的作用。 在这个项目中，我们将定义 MeCP2 对异色和常色凝聚的贡献 正常和突变神经元并分析 RTT 引起的突变对 LLPS 的影响。我们的目标是获得 深入了解 MeCP2 的功能作为设计新型治疗方法的基础。潜力新 基于这一假设的疗法需要时间才能发展成应用。探索更直接的 我们将使用表观遗传编辑作为治疗工具来激活位于的非活性 wt MECP2 等位基因 位于不活跃的 X 染色体上。最重要的是，表观遗传编辑将准确恢复 MeCP2 的表达 野生型水平，从而避免 MeCP2 过度表达的毒性后果。相比之下，其他策略 例如使用载体介导的 MeCP2 转导将总是产生过度表达 MeCP2 的细胞 因此会导致严重的副作用，如 MECP2 重复综合征患者所见。