Mechanisms of ATM activation

ATM 激活机制

• 批准号: 7590935
• 负责人: TANYA T PAULL
• 金额: $ 25.05万
• 依托单位: UNIVERSITY OF TEXAS AT AUSTIN
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-05-01 至 2014-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7590935
• 关键词: ATM Signaling Pathway; ATM activation; ATM deficient; ATM function; Acetylation; Address; Apoptosis; Ataxia Telangiectasia; Ataxia-Telangiectasia-Mutated protein kinase; Binding; Biochemical; Cell Cycle Arrest; Cell Cycle Checkpoint; Cells; Chronic; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA biosynthesis; Databases; Dimerization; Doctor of Philosophy; Eukaryotic Cell; Exhibits; Exposure to; Frequencies; Genomic Instability; Genotype; Goals; Homodimerization; Human; Hydrogen Peroxide; Hypersensitivity; In Vitro; Investigation; Ionizing radiation; Malignant Neoplasms; Mammalian Cell; Mediating; Metabolic; Molecular; Mus; Mutate; Nerve Degeneration; Oncogenic; Oxidative Stress; Pathway interactions; Patients; Phosphorylation; Population; Principal Investigator; Process; Proteins; Recombinants; Recruitment Activity; Regulation; Repair Complex; Resistance; Role; Signal Transduction; Site; System; TP53 gene; Testing; Tumor Suppressor Proteins; Work; ataxia telangiectasia mutated protein; base; biological adaptation to stress; dimer; inhibitor/antagonist; monomer; mutant; novel; oxidation; oxidative damage; prevent; programs; progressive neurodegeneration; protein protein interaction; public health relevance; reconstitution; response; sensor; tumor; tumor progression; tumorigenesis

DESCRIPTION (provided by applicant): The ATM protein kinase is a master regulator of the cellular response to chromosomal DNA double-strand breaks. This type of DNA damage occurs during DNA replication, as a result of damage from metabolic intermediates, and after exposure to ionizing radiation and radiomimetic compounds. In response to DNA damage, ATM phosphorylates many cellular substrates, several of which are known tumor suppressors in humans. Phosphorylation of these substrates initiates cell cycle arrest, apoptosis, and DNA repair. Loss of ATM, as seen in patients with Ataxia-Telangiectasia (A-T), results in increased genomic instability, a complete loss of double-strand break-induced cell cycle checkpoints, and a significant increase in cancer frequency. A-T patients also suffer from severely reduced responses to oxidative stress as well as to DNA double-strand breaks, and chronic oxidative stress has been shown to contribute to neurodegeneration seen in these patients. The ATM protein in mammalian cells exists as an inactive homodimer and becomes activated by DNA double-strand breaks through association with the DNA repair complex Mre11/Rad50/Nbs1 (MRN). In previous work we have reconstituted the ATM activation process with recombinant purified proteins and showed that MRN acts as a double-strand break sensor for ATM. In the current proposal we use this system to characterize the mechanisms of ATM activation in greater detail and investigate novel pathways of ATM regulation. Specific Aim 1 addresses the molecular basis of ATM activation through oxidative damage and seeks to identify ATM domains and residues involved in this process. Analysis of specific mutants deficient in oxidative activation will be used to investigate the functions of this pathway in human cells. Aim 2 investigates the mechanistic role of Nbs1 in ATM activation by double-strand breaks and addresses the functional relationship between MRN and ATM in greater detail through the identification of MRN-ATM protein-protein interactions and ATM homodimerization motifs. Aim 3 characterizes the roles of other proteins that are known to regulate ATM activation and substrate phosphorylation in human cells, and investigates the functional effects of ATM acetylation and autophosphorylation. The overall goal of the project is to biochemically decipher the many layers of regulation that govern ATM activity in human cells in order to understand how this important protein responds so rapidly and specifically to DNA double-strand breaks and oxidative stress. PUBLIC HEALTH RELEVANCE: The ATM protein kinase is activated by DNA damage to initiate cell cycle arrest, programmed cell death, and DNA repair. These responses are essential for preventing oncogenic transformation in humans, and loss of ATM has been shown to promote tumorigenesis. Greater understanding of the mechanisms of ATM activation is essential for our understanding of the primary cellular defense against genomic instability and tumor progression.

描述（由申请人提供）：ATM蛋白激酶是对染色体DNA双链断裂的细胞反应的主要调节剂。由于代谢中间体的损伤以及暴露于电离辐射和放射性化合物后，这种类型的DNA损伤在DNA复制过程中发生。为了应对DNA损伤，ATM磷酸化了许多细胞底物，其中一些是人类中已知的肿瘤抑制剂。这些底物的磷酸化引发了细胞周期停滞，凋亡和DNA修复。在共济失调性省telangiyctia（A-T）的患者中，ATM的损失导致基因组不稳定性增加，双链破裂引起的细胞周期检查点的完全丧失以及癌症频率的显着增加。 A-T患者还严重降低了对氧化应激以及对DNA双链断裂的反应，并且已显示慢性氧化应激有助于这些患者的神经变性。哺乳动物细胞中的ATM蛋白作为一种非活性均二聚体存在，并通过与DNA修复复合物MRE11/RAD50/NBS1（MRN）缔合而被DNA双链破裂激活。在先前的工作中，我们已经通过重组纯化的蛋白质重组了ATM激活过程，并表明MRN充当ATM的双链断裂传感器。在当前的建议中，我们使用该系统来详细介​​绍ATM激活的机制，并研究ATM调节的新型途径。特定目标1通过氧化损伤解决了ATM激活的分子基础，并试图识别此过程中涉及的ATM域和残基。分析缺乏氧化激活的特定突变体将用于研究该途径在人类细胞中的功能。 AIM 2通过双链断断裂研究了NBS1在ATM激活中的机械作用，并通过鉴定MRN-ATM蛋白 - 蛋白质相互作用和ATM均二聚化基序来更详细地解决MRN和ATM之间的功能关系。 AIM 3表征了其他已知蛋白质的作用，这些蛋白质已知，这些蛋白质在人类细胞中调节ATM激活和底物磷酸化，并研究ATM乙酰化和自磷酸化的功能作用。该项目的总体目的是在生物化学上破译人类细胞中ATM活性的许多调节层，以了解该重要蛋白如何迅速，特别是对DNA双链断裂和氧化应激的反应。公共卫生相关性：ATM蛋白激酶通过DNA损伤激活以启动细胞周期停滞，程序性细胞死亡和DNA修复。这些反应对于预防人类的致癌转化至关重要，并且已证明ATM的丧失促进肿瘤发生。对ATM激活机制的更多了解对于我们对基因组不稳定性和肿瘤进展的主要细胞防御的理解至关重要。