Connecting 3D genome misfolding to transcriptional silencing in fragile X syndrome
将 3D 基因组错误折叠与脆性 X 综合征中的转录沉默联系起来
基本信息
- 批准号:10634553
- 负责人:
- 金额:$ 43.39万
- 依托单位:
- 依托单位国家:美国
- 项目类别:
- 财政年份:2019
- 资助国家:美国
- 起止时间:2019-08-15 至 2024-06-30
- 项目状态:已结题
- 来源:
- 关键词:
项目摘要
Title: Connecting 3D genome misfolding to transcriptional silencing in fragile X syndrome
Project Summary/Abstract
More than 30 inherited neurological disorders, including fragile X syndrome (FXS), Huntington's disease,
amyotrophic lateral sclerosis, and Friedreich's ataxia, are caused by the unstable expansion of repetitive DNA
sequences termed short tandem repeats (STRs). In FXS, STR expansion above a critical length threshold
causes pathogenic silencing of the STR-containing gene, resulting in severe neuropsychiatric symptoms. An
increased understanding of the molecular mechanisms governing how STR expansion contributes to
transcriptional silencing would facilitate efforts to develop treatments for repeat expansion disorders driven by
pathologic gene expression changes. The objective of this proposal is to understand the link among higher-order
chromatin architecture, repressive chromatin modifications, architectural protein occupancy, and gene
expression silencing in FXS. Recently, we discovered that nearly all disease-associated STRs (daSTRs) are
located at boundaries demarcating 3D chromatin domains. daSTRs specifically localize to ultra-high-density CpG
island boundaries, suggesting they might be hotspots for epigenetic instability or topological disruption upon STR
expansion. Consistent with this idea, we found that FXS patients exhibit severe boundary disruption and loss of
CTCF occupancy in a manner that correlates with the degree of FMR1 silencing. Due to the boundary disruption,
the FMR1 gene undergoes a topological switch from the downstream domain containing numerous putative
enhancers to the upstream domain devoid of regulatory elements. Based on these findings, we hypothesized
that 3D genome miswiring is causally linked to pathologic silencing of FMR1 via the gene's topological switch
from an active to silenced regulatory landscape. We will test our hypothesis with three Specific Aims. First, we
will dissect the cause-and-effect link between 3D genome misfolding and FMR1 silencing. We will assay
architecture and FMR1 expression after mutating CTCF and YY1 binding sites at the FMR1 boundary with
CRISPR-Cas9 and ectopically silencing FMR1 with the dCas9-KRAB repressor in healthy cells. Second, we will
create genome-wide maps of CTCF/YY1 binding, H3K9me3/H3K27me3 repressive domains, and DNA
methylation in FXS samples with a range of CGG daSTR lengths. By computationally integrating this data, we
will elucidate how STR tract length affects distal epigenetic modifications to disrupt chromatin architecture and
FMR1 expression. Third, we will re-engineer genome topology in FXS patient cells via synthetic architectural
proteins and active de-methylation of specific architectural motifs. We will determine the degree to which 3D
genome engineering alone or in combination with linear Epigenome engineering will reprogram repressive
chromatin marks and de-repress FMR1. Our work is significant because it will shed new light into how 3-D
epigenetic mechanisms go awry and can be repaired during the acquisition and progression of
neurodevelopmental and neurodegenerative disease states.
标题:连接3D基因组与脆弱X综合征中的转录沉默错误折叠
项目摘要/摘要
30多个遗传性神经系统疾病,包括脆弱的X综合征(FXS),亨廷顿氏病,
肌萎缩性的侧面硬化症和弗里德里希(Friedreich)的共济失调是由重复性DNA的不稳定膨胀引起的
序列称为短串联重复(STR)。在FXS中,STR扩展高于临界长度阈值
引起含StR的基因的致病沉默,导致严重的神经精神症状。一个
增加对主张Str扩展如何促进的分子机制的了解
转录沉默将有助于开发以重复扩张障碍的疗法的努力
病理基因表达变化。该提案的目的是了解高阶之间的联系
染色质结构,抑制性染色质修饰,建筑蛋白占用和基因
表达沉默在FXS中。最近,我们发现几乎所有与疾病相关的STR(DASTR)是
位于边界上划定3D染色质结构域。 DASTR专门定位于超高密度CPG
岛屿边界,表明它们可能是表观遗传不稳定性或拓扑破坏的热点
扩张。与这个想法一致,我们发现FXS患者表现出严重的边界破坏和丧失
CTCF的占用率与FMR1沉默程度相关的方式。由于边界破坏,
FMR1基因从下游域进行拓扑开关,其中包含许多推定
上游领域的增强剂,没有调节元件。基于这些发现,我们假设
该3D基因组错误与通过基因的拓扑开关的病理沉默有因果关系
从积极到沉默的监管景观。我们将以三个特定目标检验我们的假设。首先,我们
将剖析3D基因组错误折叠和FMR1沉默之间的因果关系。我们将分析
在FMR1边界突变CTCF和YY1结合位点后的结构和FMR1表达
CRISPR-CAS9和异位在健康细胞中使用DCAS9-KRAB抑制剂对FMR1进行沉默。第二,我们会的
创建CTCF/YY1结合,H3K9ME3/H3K27ME3抑制域和DNA的全基因组图
具有一系列CGG DASTR长度的FXS样品中的甲基化。通过计算整合这些数据,我们
将阐明str道长度如何影响远端表观遗传学修饰,以破坏染色质结构和
FMR1表达。第三,我们将通过合成体系结构重新设计FXS患者细胞中的基因组拓扑。
蛋白质和特定结构基序的主动去甲基化。我们将确定哪个3D的程度
基因组工程单独或与线性表观基因组工程结合使用,将重新编程
染色质标记和去抑制FMR1。我们的工作很重要,因为它将为3-D带来新的启示
表观遗传机制出现问题,可以在获取和进展过程中修复
神经发育和神经退行性疾病状态。
项目成果
期刊论文数量(2)
专著数量(0)
科研奖励数量(0)
会议论文数量(0)
专利数量(0)

暂无数据
数据更新时间:2024-06-01
Jennifer Elizabeth...的其他基金
From 3D genomes to neural connectomes: Higher-order chromatin mechanisms encoding long-term memory
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Connecting 3D genome misfolding to transcriptional silencing in fragile X syndrome
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Connecting 3D genome misfolding to transcriptional silencing in fragile X syndrome
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