Role of the Rett Syndrome-causing gene MeCP2 in 3D chromosomal organization and rescue of cellular disease phenotypes

Rett 综合征致病基因 MeCP2 在 3D 染色体组织和细胞疾病表型拯救中的作用

• 批准号: 10339423
• 负责人: X. Shawn Liu
• 金额: $ 24.9万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2023-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10339423
• 关键词: 3-Dimensional; Academic Medical Centers; Affect; Alleles; Binding; Bioinformatics; Biological Assay; Brain; CCCTC-binding factor; CRISPR/Cas technology; Cell Nucleus; Cells; Cerebrum; ChIP-seq; Chromatin; Chromatin Interaction Analysis by Paired-End Tag Sequencing; Chromatin Loop; Chromosome Structures; Co-Immunoprecipitations; Collaborations; Cytosine; DNA Methylation; DNA Sequence Alteration; DNA-Binding Proteins; DNMT3a; Data; Data Set; Defect; Development; Disease; Electrophysiology (science); Enhancers; Epigenetic Process; Female; Fostering; Frameshift Mutation; Gene Expression; Gene Expression Regulation; Genes; Genetic Transcription; Genome; Genomics; Guide RNA; Health; Heterochromatin; Histone Acetylation; Histone-Lysine N-Methyltransferase; Histones; Human; Impairment; Institutes; Intellectual functioning disability; Investigation; Knock-out; Knowledge; Label; Laboratories; Laboratory Research; Lentivirus; Link; Liquid substance; Maps; Mediating; Mental Retardation; Methyl-CpG-Binding Protein 2; Microcephaly; Mission; Modification; Molecular; Morphology; Mus; Mutation; National Institute of Mental Health; Neighborhoods; Neurobiology; Neurodevelopmental Disorder; Neurons; Neurotransmitters; Nuclear Proteins; Nucleosomes; Organoids; Pathway interactions; Patients; Phase; Phenotype; Play; Property; Proteins; Protocols documentation; Psyche structure; Public Health; RNA analysis; Regulation; Reporter; Research; Research Personnel; Rett Syndrome; Role; Series; Symptoms; Syndrome; System; Testing; Therapeutic; Transcriptional Regulation; Transgenes; United States National Institutes of Health; Western Blotting; X Chromosome; X Inactivation; autism spectrum disorder; brain cell; brain size; cohesin; demethylation; disability; disease phenotype; electrical property; experimental study; high resolution imaging; human embryonic stem cell; improved; induced pluripotent stem cell; innovation; insight; loss of function mutation; male; mammalian genome; mature animal; mutant; nervous system disorder; neuronal cell body; neurophysiology; novel; novel therapeutic intervention; postnatal; precursor cell; promoter; restoration; tool; transcriptome sequencing

Modified Project Summary/Abstract Section As an NIH Pathway to Independence K99/R00 awardee, I am moving to Columbia University Medical Center to start my research laboratory as a principle investigator for the R00 phase. Rett syndrome (RTT) is an X-linked postnatal progressive neurodevelopmental disorder associated with severe mental disability and autism-like syndromes. The disease is caused by loss-of-function mutations of the DNA binding protein MeCP2 (Methyl CpG-binding Protein 2) in the X chromosome and represents the second most common cause of intellectual disability in females. Loss of MeCP2 leads to expression changes in thousands of genes, compromises the majority of brain cells and circuits, and dysregulates all neurotransmitter systems. However, how MeCP2 can act as a global repressor of gene activity as well as an activator for gene expression remains an open question in the field. Microcephaly (the reduction in brain size) has been documented as a hallmark of RTT, and analysis of hESC/iPSC-derived RTT neurons showed a reduced soma size as well. Our preliminary studies on human RTT mutant neurons showed a panel of cellular phenotypes including reduced soma size, impaired electrical properties, and defects in chromosomal structures. Therefore, we hypothesized that MeCP2 is involved in the organization of 3D chromosomal landscape contributing to the regulation of gene expression and subsequent neurobiology. We demonstrated that MeCP2 proteins form dynamic liquid-like condensates at the heterochromatic regions and concentrate heterochromatic factor HP1α but not components of active transcription in the nucleus. This condensate property of MeCp2 contributes to the compartmentation of 3D genome and the regulation of transcription machinery (Aim 1, K99 phase). Then we found that the intrinsically disordered region-2 (IDR-2) of MeCP2 protein mediates the formation of heterochromatin condensate. A common RTT mutant MeCP2-R168X lacking IDR-2 fails to form heterochromatin condensates to concentrate the heterochromatic factor and causes defects in the transcription regulation, providing a molecular mechanism of MeCP2-mediated 3D chromosomal organization (Aim 2, K99 phase). Development of RTT-like symptoms in mice can be reversed in RTT adult animals following the restoration of MeCP2 expression. As most female RTT patients still carry a wild type allele of MeCP2 subject to the random X-chromosome inactivation (XCI), it will be of therapeutic benefit if the wild type allele of MeCp2 in the inactive X chromosome (Xi) can be reactivated. We developed a DNA methylation editing tool by fusion of a catalytically inactive Cas9 with Tet1/Dnmt3a. Recently we expanded this toolbox to manipulate other epigenetic modifications including histone acetylation and DNA looping. We will use these tools to reverse the RTT phenotypes via reactivation of the wild type MECP2 allele on the Xi (Aim 3, R00 phase).This project will fill the gaps in our knowledge of MeCP2 function in the organization of 3D chromosomal structure and test the novel therapeutic approach to reverse RTT phenotypes.

修改的项目摘要/摘要部分