Molecular and neurodevelopmental consequences of ADNP mutation

ADNP 突变的分子和神经发育后果

• 批准号: 10372679
• 负责人: Kavitha Sarma
• 金额: $ 29.51万
• 依托单位: WISTAR INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-20 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10372679
• 关键词: Affect; Binding Sites; Brain; Cerebrum; Chromatin; Chromatin Structure; DNA; DNA Damage; Data; Data Set; Defect; Deletion Mutation; Development; Disease; Etiology; Exhibits; Fragile X Syndrome; Functional disorder; Future; Gene Expression; Gene Expression Profile; Genes; Genetic Transcription; Genome; Genomic Segment; Homeobox; Human; Hybrids; Label; Link; Malignant Neoplasms; Mediating; Modeling; Molecular; Morphology; Mus; Mutate; Mutation; Neurodegenerative Disorders; Neurodevelopmental Disorder; Neuronal Differentiation; Neurons; Oncogene Deregulation; Organoids; Population; Proteins; Proteomics; RNA; Reporting; Research; Ribonuclease H; Single-Stranded DNA; Site; Structure; Syndrome; Therapeutic Intervention; Tyrosine; Undifferentiated; attenuation; autism spectrum disorder; base; chromatin remodeling; embryonic stem cell; frontotemporal lobar dementia-amyotrophic lateral sclerosis; genetic regulatory protein; genome-wide; helicase; homeodomain; induced pluripotent stem cell; insight; mutant; nerve stem cell; new therapeutic target; nucleic acid structure; progenitor; programs; single-cell RNA sequencing; transcription factor

PROJECT SUMMARY The abundance of mutations in chromatin regulatory proteins in autism spectrum disorders (ASD) highlights the significance of chromatin structure in normal brain development. A relatively new and little understood chromatin structure that has been implicated in several neurodevelopmental disorders is the R-loop. R-loops are three- stranded, nucleic acid structures containing a DNA:RNA hybrid and a displaced single stranded DNA, that form and resolve continually. However, when R-loops accumulate, they can impede gene transcription, promote DNA damage, and result in disease. R-loops are implicated in several neurodegenerative disorders, such as frontotemporal dementia and amyotrophic lateral sclerosis, and in neurodevelopmental disorders such as Fragile X Syndrome. Whether R-loop deregulation contributes to the etiology of ASD is not clear. The central challenge in linking R-loop deregulation to ASDs has been pinpointing factors that are mutated in ASD that are also R-loop regulators. We have identified factors that localize to R-loops using a proximity labeling proteomic approach. Our approach yielded chromatin remodelers, homeobox transcription factors, and helicases. A number of these proteins had also been reported to be mutated in ASD, e.g. ADNP, POGZ, CHD2, and DHX30. These preliminary results provide the first indication that R-loop dysfunction may contribute to ASD. We focus our study on the analysis of ADNP, an R-loop regulator that is frequently mutated in ASD and is causal in ADNP syndrome. Our preliminary studies show that ADNP loss in mouse embryonic stem cells (mESCs) results in R-loop accumulation specifically at ADNP binding sites. Importantly, our data show that the homeodomain of ADNP is critical for R-loop suppression and that deletion of the homeodomain compromises the ability of mESCs to differentiate into neural progenitor cells (NPCs). We found that human induced pluripotent stem cells (hiPSCs) with an ADNP tyrosine 719* mutation (ADNP Tyr719*) also show R-loop deregulation. Based on these data, we hypothesize that accumulation of R-loops at specific genomic regions changes normal gene expression programs leading to improper neuronal differentiation, which can ultimately contribute to ASD. Here we will determine how R-loop accumulation in ADNP Tyr719* hiPSCs affects gene expression in the pluripotent state and upon neuronal differentiation. We will evaluate the neurodifferentiation potential of ADNP Tyr719* using a cerebral organoid model and determine if R-loop attenuation can ameliorate the neurodifferentiation defects observed in ADNP mutants.

项目概要 自闭症谱系障碍 (ASD) 中染色质调节蛋白的大量突变凸显了 染色质结构在正常大脑发育中的重要性。一种相对较新且知之甚少的染色质 R 环是与多种神经发育障碍有关的结构。 R 环是三- 含有 DNA:RNA 杂交体和置换的单链 DNA 的链状核酸结构，形成 并不断地解决。然而，当 R 环积累时，它们会阻碍基因转录，促进 DNA 损害，并导致疾病。 R 环与多种神经退行性疾病有关，例如 额颞叶痴呆和肌萎缩侧索硬化症，以及神经发育障碍，例如脆性 X综合症。 R 环放松管制是否会导致自闭症谱系障碍 (ASD) 的病因尚不清楚。核心挑战 将 R 环放松管制与自闭症谱系障碍 (ASD) 联系起来，已查明自闭症谱系障碍 (ASD) 中突变的因素也是 R 环 监管机构。我们使用邻近标记蛋白质组学方法确定了定位于 R 环的因子。 我们的方法产生了染色质重塑剂、同源框转录因子和解旋酶。其中一些 据报道，自闭症谱系障碍中的蛋白质也发生了突变，例如ADNP、POGZ、CHD2 和 DHX30。这些 初步结果首次表明 R 环功能障碍可能导致自闭症谱系障碍 (ASD)。我们专注于学习 ADNP 是一种 R 环调节因子，在 ASD 中经常发生突变，并且与 ADNP 综合征有关。 我们的初步研究表明，小鼠胚胎干细胞 (mESC) 中 ADNP 缺失会导致 R 环 特异性地在 ADNP 结合位点积累。重要的是，我们的数据表明 ADNP 的同源域是 对于 R 环抑制至关重要，并且同源域的删除会损害 mESC 的能力 分化为神经祖细胞（NPC）。我们发现人类诱导多能干细胞（hiPSC） 具有 ADNP 酪氨酸 719* 突变 (ADNP Tyr719*) 也表现出 R 环失调。根据这些数据，我们 假设 R 环在特定基因组区域的积累会改变正常的基因表达 导致神经元分化不当的程序，最终可能导致自闭症谱系障碍（ASD）。在这里我们将 确定 ADNP Tyr719* hiPSC 中 R 环积累如何影响多能状态下的基因表达 以及神经元分化。我们将使用 ADNP Tyr719* 的神经分化潜力进行评估 脑类器官模型并确定 R 环衰减是否可以改善神经分化缺陷 在 ADNP 突变体中观察到。