Trinucleotide repeat disorders and the 3D genome

三核苷酸重复紊乱和 3D 基因组

• 批准号: 10251290
• 负责人: Linda Zhou
• 金额: $ 2.57万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-09-01 至 2022-05-16
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10251290
• 关键词: 3-Dimensional; Ablation; Address; Affect; Alleles; Anxiety; Architecture; Binding; Cataract; Cell Line; Cells; Chromatin; Complex; DNA Sequence; Data; Defect; Dimensions; Disease; Endonuclease I; Engineering; Epigenetic Process; Evolution; FMR1; Fragile X Syndrome; Friedreich Ataxia; Functional disorder; Gene Expression; Genes; Genetic; Genetic Transcription; Genome; Genome engineering; Heart Abnormalities; Hi-C; Human; Hyperactivity; Individual; Inherited; Lead; Length; Light; Link; Location; Maps; Measures; Mediating; Molecular; Mutate; Myotonic Dystrophy; Outcome; Patients; Phenotype; Quantitative Reverse Transcriptase PCR; Repetitive Sequence; Resolution; Role; Seizures; Series; Severity of illness; Short Tandem Repeat; Site; Structure; Sum; Symptoms; Testing; Therapeutic Intervention; Trinucleotide Repeat Expansion; Trinucleotide Repeats; Work; attenuation; base; causal variant; experimental study; genome editing; induced pluripotent stem cell; insight; novel therapeutic intervention; prevent; recruit; respiratory; social deficits; targeted treatment; therapeutic target; therapy development

Project Summary Unstable expansion of repetitive DNA sequences termed short tandem repeats (STRs) serves as the mechanistic basis for more than 25 inherited human disorders. Patients with unstable repeat expansion diseases suffer from a complex array of symptoms, including: cardiac defects, cataracts, anxiety, hyperactivity, low IQ, social deficits, respiratory defects and seizures. In some diseases, such as Fragile X Syndrome and Freidreich’s Ataxia, the downstream phenotype is mediated in large part by reduced gene expression. In all of these diseases, continuous repeat expansion is associated with disease severity. Treating trinucleotide repeat disorders is thus complex because the primary effectors of disease include both the continuous expansion of repetitive sequences as well as disrupted expression of the gene containing the repeat. Thus, an increased understanding of the molecular mechanisms governing STR instability and expansion related gene dysregulation would facilitate efforts to develop therapies to prevent and treat repeat expansion disorders. In our preliminary work, we introduce the higher order chromatin architecture as a new dimension in understanding these features in repeat expansion disorders. Our data shows that (1) the large majority of disease associated STRs are located precisely at boundaries demarcating 3D genome folding domains termed topologically associating domains (TADs) and subTADs and (2) repeat expansion in the FMR1 gene, the genetic driver for Fragile X Syndrome, results in CTCF occupancy ablation and large-scale TAD/subTAD reorganization in a manner that correlates with STR tract length, disease severity, and transcriptional disruption of FMR1. Given the increasing importance of the 3D genome, there is a critical need to extend our preliminary data to understand how the 3D genome may be perturbed by trinucleotide repeat expansion and whether this perturbation could contribute to the primary effectors of disease originating from the causal gene itself: repeat instability and dysregulated gene expression. This proposal outlines the next steps doing so. In the first aim, I will perform genome engineering experiments to determine if the domain reorganization we have observed around FMR1 contributes to decreased gene expression. In the second aim, I will create high resolution topological maps around the FXN gene, the genetic driver for Freidreich’s ataxia, the determine whether boundary disruption is present in an additional trinucleotide repeat disorder. In the third aim, I will perform additional genome editing experiments to elucidate whether domain boundary disruptions can influence repeat instability and gene expression of the FXN gene. In sum, the accomplishment of these aims would demonstrate that the 3D genome can be perturbed in repeat expansion disorders and that this perturbation can mediate repeat instability and disrupted gene expression. Ultimately, we could use these results to determine whether manipulating the 3D genome could be a potential therapeutic target for treating these diseases.

项目概要 称为短串联重复 (STR) 的重复 DNA 序列的不稳定扩展可作为 超过 25 种人类遗传性疾病的机制基础。重复扩增不稳定的患者 疾病会带来一系列复杂的症状，包括：心脏缺陷、白内障、焦虑、多动、 智商低、社交缺陷、呼吸系统缺陷和癫痫发作。在某些疾病中，例如脆性 X 综合征和 弗赖德赖希共济失调，下游表型在很大程度上是由基因表达减少介导的。在所有的 这些疾病的持续重复扩张与疾病的严重程度相关。处理三核苷酸重复 因此，疾病是复杂的，因为疾病的主要效应器包括 重复序列以及含有重复序列的基因的表达被破坏。因此，增加了 了解控制 STR 不稳定性和扩增相关基因的分子机制 失调将有助于开发预防和治疗重复扩张障碍的疗法。 在我们的前期工作中，我们引入了高阶染色质结构作为一个新的维度 了解重复扩张障碍的这些特征。我们的数据表明（1）绝大多数疾病 相关的 STR 精确地位于划分 3D 基因组折叠域的边界处，称为 拓扑关联结构域 (TAD) 和 subTAD，以及 (2) FMR1 基因中的重复扩展，即遗传性 脆性 X 综合征的驱动因素，导致 CTCF 占用消融和大规模 TAD/subTAD 重组 其方式与 STR 束长度、疾病严重程度和 FMR1 转录破坏相关。 鉴于 3D 基因组的重要性日益增加，迫切需要将我们的初步数据扩展到 了解 3D 基因组如何受到三核苷酸重复扩展的干扰，以及这是否 扰动可能导致源自致病基因本身的疾病的主要效应：重复 不稳定和失调的基因表达。该提案概述了这样做的后续步骤。在第一个目标中，我 将进行基因组工程实验以确定我们是否观察到了域重组 FMR1 周围的区域导致基因表达降低。第二个目标是创造高分辨率 围绕 FXN 基因的拓扑图，Freidreich 共济失调的遗传驱动因素，确定是否 边界破坏存在于额外的三核苷酸重复紊乱中。在第三个目标中，我将实现 额外的基因组编辑实验，以阐明域边界破坏是否会影响重复 FXN 基因的不稳定性和基因表达。总而言之，这些目标的实现将表明 3D 基因组可能会在重复扩张紊乱中受到干扰，并且这种干扰可以介导 重复不稳定并破坏基因表达。最终，我们可以使用这些结果来确定是否 操纵 3D 基因组可能是治疗这些疾病的潜在治疗靶点。