Base Excision DNA Repair in Premature Aging and Neurodegeneration

过早衰老和神经退行性疾病中的碱基切除 DNA 修复

• 批准号: 8736611
• 负责人: David Wilson
• 金额: $ 41.2万
• 依托单位: NATIONAL INSTITUTE ON AGING
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8736611
• 关键词: APTX gene; Aging; Aging-Related Process; Ataxia; Atrophic; Autophagocytosis; Base Excision Repairs; Base Sequence; Biochemical; Biological; Biological Assay; Brain; CAG repeat; Cell physiology; Cells; Cerebellum; Cessation of life; Cockayne Syndrome; Collaborations; Complex; Corpus striatum structure; Cutaneous; DNA; DNA Damage; DNA Excision Repair Protein ERCC-6; DNA Modification Process; DNA Repair; DNA Sequence; DNA glycosylase; DNA-Directed RNA Polymerase; Data; Defect; Development; Disease; ERCC6 gene; Exhibits; Eye; Failure; Free Radicals; Future; Genes; Growth; Hereditary Disease; Human; Huntington Disease; In Vitro; Inborn Genetic Diseases; Incidence; Inner mitochondrial membrane; Lesion; Ligation; Longevity; Malignant Neoplasms; Mitochondria; Mitochondrial DNA; Mitochondrial RNA; Modeling; Molecular; Motor; Mutation; Myotonic Dystrophy; Nerve Degeneration; Neurologic; Neurons; Nuclear; Nucleic Acids; Organism; Outcome; Oxidative Stress; Pathology; Pathway interactions; Patients; Peripheral; Phenotype; Phosphoric Monoester Hydrolases; Photosensitivity; Play; Polynucleotide 5' -Hydroxyl-Kinase; Positioning Attribute; Predisposition; Premature aging syndrome; Process; Production; Property; Proteins; Purkinje Cells; Reaction; Reactive Oxygen Species; Recruitment Activity; Repair Complex; Reporting; Role; Single Strand Break Repair; Source; Stress; Symptoms; Tissues; Toxic effect; Transcript; Work; adenylate; age related; base; coping; crosslink; disease phenotype; genetic linkage; human APEX1 protein; insight; mitochondrial autophagy; mitochondrial dysfunction; mitochondrial genome; nerve stem cell; nervous system disorder; oculomotor; oxidative DNA damage; oxidative damage; repaired; response; sensor; sensory neuropathy; stem; stoichiometry; theories; tyrosyl-DNA phosphodiesterase

Expansion of CAG/CTG repeats in DNA is the underlying cause of >14 genetic disorders, including Huntington disease (HD) and myotonic dystrophy. The mutational process is ongoing, with increases in repeat size enhancing the toxicity of the expansion in specific tissues. In many repeat diseases, the repeats exhibit high instability in the striatum, whereas instability is minimal in the cerebellum. In recent work, we have provided molecular insights into how BER protein stoichiometry may contribute to the tissue-selective instability of CAG/CTG repeats by using specific repair assays. In particular, repair efficiency at CAG/CTG repeats and at control DNA sequences was markedly reduced under conditions that mimic the striatal situation, likely because of lower levels of the proteins APE1, FEN1, and LIG1. Moreover, damage located toward the 5' end of the repeat tract was poorly repaired, with the accumulation of incompletely processed intermediates as compared to an abasic lesion in the center or at the 3' end of the repeats or within control sequences. In addition, repair of lesions at the 5' end of CAG or CTG repeats involved multinucleotide synthesis, particularly at the cerebellar stoichiometry, suggesting that long-patch BER processes lesions at sequences susceptible to hairpin formation. Our results show that the BER stoichiometry, nucleotide sequence, and DNA damage position modulate repair outcome and suggest that a suboptimal long-patch BER activity promotes CAG/CTG repeat instability. More recently, we have found that the BER DNA glycosylase NEIL1 contributes to germline and somatic CAG repeat expansion in HD. Single-strand break repair (SSBR) is an important subpathway of BER. Recent data has found a genetic linkage between proteins of SSBR aprataxin, tyrosyl-DNA phosphodiesterase 1 and DNA polynucleotide kinase phosphatase and human neurological disorders, implicating this process in protection against neuronal cell loss and brain function. Ataxia with oculomotor apraxia 1 (AOA1) is caused by mutation in the APTX gene, which encodes the stand break repair protein aprataxin. Aprataxin removes 5-adenylate groups in DNA that arise from aborted ligation reactions. AOA1 is characterized by global cerebellar atrophy, highlighted by loss of Purkinje cells, ocular motor apraxia, and motor and sensory neuropathy. Strikingly, AOA1 patients lack the cancer susceptibility and other peripheral symptoms (e.g., immunological deficiencies) commonly associated with other inherited disorders stemming from a DNA repair defect. We have reported that aprataxin activity is indispensable for maintaining mitochondrial function, indicating that there is likely a mitochondrial component to the disease phenotype of AOA1. Moreover, our data indicate that because of their higher BER capacity, proliferative neural progenitor cells are more efficient at repairing DNA damage compared with their neuronally differentiated progeny. Future studies are aimed at determining the reason behind the tissue selectivity of AOA1, with an eye towards differential repair capacity as a key factor. Cockayne Syndrome (CS) is an autosomal recessive disorder, characterized by growth failure, neurological abnormalities, premature aging symptoms, and cutaneous photosensitivity, but no increased cancer incidence. CS is divided into two strict complementation groups: CSA (mutation in CKN1) and CSB (mutation in ERCC6). Of the patients suffering from CS, 80% have mutations in the CSB gene. We are pursuing the hypothesis that the primary role of CS proteins is to facilitate the repair of endogenous DNA damage, and we have evidence for a direct role of CSB in regulating BER efficiency. Our in vitro work has also helped define the biochemical properties of CSB, revealing that the protein interacts with a diverse range of nucleic acid substrates and likely has important ATP-dependent and ATP-independent functions. More recent results, obtained in collaboration with Dr. Vilhelm Bohr, suggest that CSB plays a direct role in not only nuclear BER, but in mitochondrial BER, likely by helping recruit, stabilize, and/or retain BER proteins in repair complexes associated with the inner mitochondrial membrane. Moreover, CSB appears to act as a mitochondrial DNA damage sensor, inducing mitochondrial autophagy in response to stress, and thus, pharmacological modulators of autophagy are potential treatment options for this accelerated aging phenotype. CSB-deficient cells also exhibit a defect in efficient mitochondrial transcript production and the CSB protein specifically promotes elongation by the mitochondrial RNA polymerase suggesting that the pathologies associated with CS are in part, a direct result of the roles that CSB plays in mitochondria. Future work will aim to determine the biological substrates and molecular role(s) of the CS proteins in endogenous DNA damage repair.

DNA中CAG/CTG重复序列的扩展是> 14种遗传疾病的根本原因，包括亨廷顿疾病（HD）和肌发育不良。 突变过程正在进行中，重复大小的增加增强了特定组织扩张的毒性。 在许多重复疾病中，重复症在纹状体中表现出很高的不稳定性，而小脑中的不稳定最小。 在最近的工作中，我们为BER蛋白化学计量法可能如何通过使用特定的维修测定法提供了分子见解。 特别是，在模仿纹状体情况的条件下，CAG/CTG重复序列和对照DNA序列的修复效率显着降低，这可能是由于蛋白质APE1，FEN1和LIG1的水平较低。 此外，与中心的无碱性病变或重复序列的3'末端相比，位于重复道5'末端的损伤的修复程度不佳，未完全处理的中间体的积累。 此外，在CAG或CTG重复量的5'末端修复病变涉及多核苷酸合成，尤其是在小脑化学计量学上，这表明长块BER在易于发夹形成的序列中的长期ber过程病变。 我们的结果表明，BER化学计量，核苷酸序列和DNA损伤位置调节修复结果，并表明次优的长距离BER活性促进了CAG/CTG重复不稳定性。 最近，我们发现BER DNA糖基酶Neil1有助于HD中的种系和体细胞CAG重复膨胀。 单链断裂修复（SSBR）是BER的重要子路口。最近的数据发现，SSBR Aprataxin，酪酶-DNA磷酸二酯酶1和DNA多核苷酸激酶磷酸酶的蛋白质与人类神经系统疾病之间存在遗传联系，这暗示了这种过程在保护神经元细胞损失和脑功能上。 用眼动替型1（AOA1）的共济失调是由APTX基因突变引起的，该基因编码了支架破裂蛋白aprataxin。 Aprataxin在DNA中除去了由中止的连接反应引起的5-腺苷酸基。 AOA1的特征是全球小脑萎缩，以purkinje细胞，眼运动失调以及运动和感觉神经病的损失突出。 令人惊讶的是，AOA1患者缺乏癌症的敏感性和其他与其他因DNA修复缺陷引起的遗传性疾病相关的外周症状（例如免疫缺陷）。 我们报道说，Aprataxin活性对于维持线粒体功能是必不可少的，表明AOA1疾病表型可能存在线粒体成分。 此外，我们的数据表明，由于其较高的BER能力，与神经元分化的后代相比，增生性神经祖细胞在修复DNA损伤方面更有效。 未来的研究旨在确定AOA1的组织选择性背后的原因，以差异修复能力为关键因素。 Cockayne综合征（CS）是一种常染色体隐性疾病，其特征是生长衰竭，神经系统异常，过早衰老症状和皮肤光敏性，但癌症发病率没有增加。 CS分为两个严格的补体组：CSA（CKN1中的突变）和CSB（ERCC6中的突变）。 在患有CS的患者中，有80％的患者患有CSB基因突变。 我们追求的假设是，CS蛋白的主要作用是促进内源性DNA损伤的修复，并且我们有证据表明CSB在调节BER效率方面的直接作用。 我们的体外工作还有助于定义了CSB的生化特性，表明该蛋白质与多种核酸底物相互作用，并且可能具有重要的ATP依赖性和非ATP依赖性功能。 与Vilhelm Bohr博士合作获得的最新结果表明，CSB不仅在核BER中起着直接的作用，而且在线粒体BER中起着直接的作用，这可能是通过帮助募集，稳定和/或保留与内部线粒体膜相关的维修复合物中的招募，稳定和/或保留BER蛋白。 此外，CSB似乎充当线粒体DNA损伤传感器，诱导线粒体自噬会响应压力，因此，自噬的药理学调节剂是这种加速衰老表型的潜在治疗选择。 CSB缺陷型细胞还表现出有效的线粒体转录产生的缺陷，CSB蛋白特异性地通过线粒体RNA聚合酶促进延伸，这表明与CS相关的病理学是部分的直接结果，是CSB在线粒体中扮演的作用的直接结果。 未来的工作将旨在确定CS蛋白在内源性DNA损伤修复中的生物底物和分子作用。