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 蛋白激酶是细胞对染色体 DNA 双链断裂反应的主要调节因子。这种类型的 DNA 损伤发生在 DNA 复制过程中，是代谢中间体损伤的结果，以及暴露于电离辐射和拟放射化合物之后。为了响应 DNA 损伤，ATM 磷酸化许多细胞底物，其中一些是已知的人类肿瘤抑制因子。这些底物的磷酸化启动细胞周期停滞、细胞凋亡和 DNA 修复。共济失调毛细血管扩张症 (A-T) 患者中 ATM 的缺失会导致基因组不稳定性增加、双链断裂诱导的细胞周期检查点完全缺失以及癌症发生率显着增加。 A-T 患者对氧化应激和 DNA 双链断裂的反应也严重减弱，慢性氧化应激已被证明会导致这些患者的神经退行性变。哺乳动物细胞中的 ATM 蛋白以无活性的同二聚体形式存在，并通过与 DNA 修复复合物 Mre11/Rad50/Nbs1 (MRN) 结合而被 DNA 双链断裂激活。在之前的工作中，我们用重组纯化蛋白重建了 ATM 激活过程，并表明 MRN 作为 ATM 的双链断裂传感器。在当前的提案中，我们使用该系统更详细地描述 ATM 激活机制并研究 ATM 调节的新途径。具体目标 1 解决通过氧化损伤激活 ATM 的分子基础，并寻求识别参与该过程的 ATM 结构域和残基。对氧化激活缺陷的特定突变体的分析将用于研究该途径在人类细胞中的功能。目标 2 研究 Nbs1 通过双链断裂在 ATM 激活中的机制作用，并通过鉴定 MRN-ATM 蛋白质-蛋白质相互作用和 ATM 同二聚化基序更详细地探讨 MRN 和 ATM 之间的功能关系。目标 3 描述了已知调节人类细胞中 ATM 激活和底物磷酸化的其他蛋白质的作用，并研究了 ATM 乙酰化和自磷酸化的功能影响。该项目的总体目标是通过生化方式破译人类细胞中控制 ATM 活性的多层调控，以了解这种重要的蛋白质如何如此快速且特异地对 DNA 双链断裂和氧化应激作出反应。公共健康相关性：ATM 蛋白激酶由 DNA 损伤激活，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致癌转化至关重要，并且 ATM 的缺失已被证明会促进肿瘤发生。更好地了解 ATM 激活机制对于我们了解针对基因组不稳定性和肿瘤进展的主要细胞防御至关重要。