Origins of DNA damage driving pathology in human neurodegeneration

DNA损伤驱动人类神经变性病理学的起源

基本信息

批准号：
10569616
负责人：
TANYA T PAULL
金额：
$ 38.95万
依托单位：
UNIVERSITY OF TEXAS AT AUSTIN
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-02-10 至 2027-01-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10569616
关键词：
ATM deficient ATM function Active Sites Adolescence Affect Age Alleles Amyloid Fibrils Ataxia Ataxia Telangiectasia Ataxia Telangiectasia Patients Automobile Driving Base Excision Repairs Biochemistry Brain Cell Cycle Arrest Cell Line Cell model Cells Cerebellar Ataxia Cerebellum Childhood Clinical Complex Cytosine DNA DNA Damage DNA Double Strand Break DNA Repair DNA Sequence Alteration DNA Single Strand Break DNA metabolism Defect Detergents Disease Disease Progression Enzymes Event Exhibits Genes Genetic Transcription Genomic DNA Genomic Instability Health Human Hybrids In Vitro Label Lesion Location Malignant Neoplasms Maps Mass Spectrum Analysis Methylation Modeling Molecular Mutation Nerve Degeneration Neurologic Dysfunctions Neuronal Differentiation Neurons Nucleotide Excision Repair Oxidative Stress Pathologic Pathology Pathway interactions Patients Phenotype Phosphorylation Phosphotransferases Play Poly Adenosine Diphosphate Ribose Poly(ADP-ribose) Polymerases Polymers Population Proteins RNA Recombinant Proteins Regulation Resistance Role Signal Transduction Single Strand Break Repair Site Source Stress Syndrome Testing Tissue Sample Work ataxia telangiectasia mutated protein ataxia-telangiectasia like disorder biological adaptation to stress brain tissue cell type early onset experimental study genome-wide human disease motor control mutant neuron loss novel oxidation oxidative damage polymerization polypeptide progressive neurodegeneration protein aggregation protein purification proteostasis response tumor progression

项目摘要

Loss or mutations in both alleles of the gene encoding ataxia-telangiectasia mutated (ATM) kinase results in early-onset cerebellar ataxia and progressive neurodegeneration in humans. Mechanistic explanations for this phenotype, as well as for the related A-T like Disorder (ATLD) caused by rare mutations in the MRE11 gene, have been elusive despite years of work on these enzymes. ATM is a master regulator of the DNA damage response, controlling checkpoint responses and survival of DNA damage in all cell types. We have previously characterized the ATM protein kinase in vitro, using purified proteins to determine that ATM is activated at sites of DNA double-strand breaks and can also be activated independently of breaks by oxidative stress. In recent work we found that loss of ATM kinase activity results in the formation of protein aggregates—detergent-resistant insoluble forms of proteins—enriched for polypeptides with intrinsically disordered domains. These aggregates form in response to hyperactivation of poly-ADP-ribose polymerases (PARPs) that are activated at sites of transcriptional stress. Analysis of 21 patient cerebellum tissue samples also showed massive levels of aggregates as well as hyperPARylation in comparison to controls, consistent with these observations. Based on this evidence we propose that protein aggregation may play a causal role in the neurodegeneration that occurs in this disorder, similar to other forms of cerebellar ataxia and to more common late-onset neurodegeneration in the human population. Here we propose to characterize the origin of single-strand DNA breaks that occur in the absence of ATM function to test the hypothesis that the combined effects of oxidative stress and transcription-dependent damage is responsible for the strand breaks and resulting protein aggregates that are observed with loss of ATM in human neurons. We will also characterize the locations and requirements for strand breaks seen in neurons expressing ATLD alleles of MRE11 and test the hypothesis that the Mre11-Rad50-Nbs1 (MRN) complex promotes single-strand break repair using in vitro biochemistry with purified proteins. These experiments will test novel hypotheses about the functions of ATM and MRN in neurons and the origins of DNA damage during cerebellar neurodegeneration.

编码共济失调 - 舌促性突变的基因的两个等位基因的损失或突变（atm）激酶会导致早期发作的小脑共济失调和人类进行性神经退行性。该表型以及相关A-T类疾病（ATLD）的机械解释由MRE11基因中罕见的突变引起酶。 ATM是DNA损伤响应的主要调节器，控制检查点所有细胞类型中DNA损伤的反应和存活。我们以前已经表征了 ATM蛋白激酶在体外，使用纯化的蛋白质确定ATM在 DNA双链断裂，也可以通过氧化而独立于断裂来激活压力。在最近的工作中，我们发现ATM激酶活性的损失导致形成蛋白质聚集体 - 抗多蛋白的耐药性形式 - 富含多肽具有本质上无序的域。这些聚集体是为了响应于在转录应力部位激活的聚ADP-核糖聚合酶（PARP）。对21例患者小脑组织样品的分析也显示了大量的聚集体与对照组相比，与这些观察结果一致。基于在这个证据上，我们建议蛋白质聚集在在这种疾病中发生的神经变性，类似于其他形式的小脑共济失调和人口中更常见的晚期神经退行性。在这里我们建议表征在没有ATM功能的情况下发生的单链DNA断裂的起源检验氧化应激和转录依赖性综合作用的假设损坏是造成观察到的链破裂和产生的蛋白质聚集体的原因在人类神经元中失去ATM。我们还将表征位置和要求对于表达MRE11的ATLD等位基因并检验假设的神经元中的链断裂 MRE11-RAD50-NBS1（MRN）配合纯化蛋白质的生物化学。这些实验将检验有关有关的新假设在小脑期间，神经元中ATM和MRN的功能以及DNA损伤的起源神经变性。