Genome-wide dysregulation of R-loops in Ataxia Telangiectasia neurological pathogenesis

共济失调毛细血管扩张症神经发病机制中 R 环的全基因组失调

• 批准号: 10607414
• 负责人: Katherine R. Westover
• 金额: $ 4.77万
• 依托单位: EMORY UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-13 至 2026-01-12
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10607414
• 关键词: ATAC-seq; ATM Gene Mutation; ATM deficient; Affect; Age; Ataxia Telangiectasia; Ataxia Telangiectasia Patients; Autopsy; Cell Nucleus; Cell physiology; Cells; Central Nervous System; Chromatin; Clinical; Cytoplasm; DNA; DNA Damage; DNA Repair; DNA Structure; Data; Defect; Deposition; Disease; Double Strand Break Repair; Eye; Failure; Gene Expression; Genome; Goals; Hybrids; Immune; Impairment; Malignant Neoplasms; Maps; Mediating; Messenger RNA; Methyltransferase; Modeling; Modification; Molecular; Monitor; Motor; Motor Neurons; Mus; Mutation; Neurodegenerative Disorders; Neurologic; Neuronal Differentiation; Neurons; Onset of illness; Pathogenesis; Pathogenicity; Pathology; Pathway interactions; Patients; Persons; Phosphorylation; Phosphotransferases; Play; Post-Transcriptional Regulation; Protein-Serine-Threonine Kinases; Proteins; Purkinje Cells; RNA; RNA Splicing; RNA immunoprecipitation sequencing; Regulation; Role; Signal Pathway; Signal Transduction; Structure; Telangiectasis; Testing; Threonine; Transcriptional Regulation; Translating; ataxia telangiectasia mutated protein; autosome; brain tissue; cell type; clinically relevant; data integration; early childhood; genome editing; genome-wide; improved; induced pluripotent stem cell; loss of function; mRNA Translation; mental development; mimetics; motor neuron development; mutation correction; nervous system disorder; new therapeutic target; repaired; response; transcriptome sequencing; trend

Project Abstract: Ataxia Telangiectasia (AT), a multisystemic neurodegenerative disease characterized by decreasing motor coordination, mental development, immune defects, and telangiectasia of the eyes, affects up to 1 in 40,000 to 100,000 people worldwide. A recessive early childhood onset disorder, AT is caused by mutations within the ataxia telangiectasia mutated (ATM) threonine/serine kinase which plays crucial roles within the DNA damage response (DDR). However, the precise molecular mechanisms underlying AT pathogenesis and how ATM loss- of-function leads to deficient DDR remain elusive. R-loops, three stranded RNA-DNA structures composed of an DNA-RNA hybrid and a non-template DNA strand, have emerged as key components of double strand break (DSB)-induced DDR. Mounting evidence has documented critical roles of R-loops in both causing and responding to DSBs. As DSBs and the failure of their repair play major roles in the pathology of AT, R-loop dysregulation is likely to contribute to AT pathogenesis. One recently identified kinase substrate of ATM is methyltransferase like 3 (METTL3) protein, a N6-methyladenosine (m6A) methyltransferase. m6A on the RNA strand of R-loops is present inside nuclei and affects R-loop formation during DSB repair. The relationship between ATM-METTL3 phosphorylation in response to DNA damage and regulation of R-loop formation, which could play crucial roles in AT pathogenesis, has yet to be defined. Our preliminary data has demonstrated a global trend of R-loops decreasing in AT patient-derived neurons compared to healthy controls. ~20% of these lost loci were rescued in an isogenic line where the ATM mutation had been corrected. We hypothesize that in AT, the lack of METTL3 phosphorylation by ATM could globally dysregulate R-loop formation and underly AT progression. In Aim 1 we will investigate the global landscape of R-loops and analyze their effect on gene expression and chromatin accessibility throughout neuronal differentiation in healthy, AT-derived, and isogenic neurons. In Aim 2, we will define how ATM-mediated phosphorylation of METTL3 impacts the formation of R- loops. We will generate iPSC-derived motor neurons from age-matched healthy controls, AT patients, and their isogenic lines with the pathogenic mutations corrected by genome editing to systematically identify critical R- loop loci that are associated with AT and mechanistically explore the role of ATM truncations in AT progression through METTL3-dependent R-loop regulation.

项目摘要： teangicycia（AT）是一种多系统神经退行性疾病，其特征是降低运动 眼睛的协调，心理发展，免疫缺陷和毛细血管扩张，最多可影响40,000分之一 全球100,000人。 AT是一个隐性的幼儿发作障碍，是由突变引起的 催化性毛细血管炎突变（ATM）苏氨酸/丝氨酸激酶，在DNA损伤中起着至关重要的作用 响应（DDR）。然而，发病机理上的确切分子机制以及ATM损失如何 功能导致DDR缺乏仍然难以捉摸。 R环，三个链的RNA-DNA结构，由A组成 DNA-RNA混合动力车和非模板DNA链已成为双链断裂的关键组成部分 （DSB）诱导的DDR。越来越多的证据记录了R环在引起和 响应DSB。随着DSB及其维修的失败在AT的病理中起主要作用 失调可能会导致在发病机理上。最近确定的一个ATM的激酶底物是 甲基转移酶（例如3（mettl3）蛋白，一种N6-甲基腺苷（M6A）甲基转移酶。 RNA上的M6a R-loops的线存在于核内，并影响DSB修复过程中的R环形成。关系 响应DNA损伤和R环形成调节的ATM-METTL3磷酸化之间 可以在发病机理中扮演至关重要的角色，尚未定义。我们的初步数据证明了 与健康对照相比，R-loops在患者衍生的神经元中的全球趋势降低。其中约有20％ 丢失的基因座是在校正ATM突变的同基因线中救出的。我们假设在 AT，ATM缺乏METTL3磷酸化可能会在全球范围内失调的R-loop形成和基础 进展。在AIM 1中，我们将研究R环的全球景观，并分析其对基因的影响 整个神经元分化的表达和染色质的可及性 神经元。在AIM 2中，我们将定义ATM介导的METTL3的磷酸化如何影响R-的形成 循环。我们将从患者及其年龄匹配的健康对照中产生IPSC衍生的运动神经元 通过基因组编辑纠正的致病突变的等源性线，系统地识别关键的R- 与AT和机械机械探索ATM截断在AT进程中的作用的循环基因座 通过依赖METTL3的R环调节。