Coordination of DNA repair and transcription by ubiquitin modification at DNA double strand breaks

DNA 双链断裂处泛素修饰协调 DNA 修复和转录

• 批准号: 10599965
• 负责人: Bin Wang
• 金额: $ 36.32万
• 依托单位: UNIVERSITY OF TX MD ANDERSON CAN CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-04-01 至 2025-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10599965
• 关键词: BRCA1 gene; Cells; Characteristics; Chromatin; Complex; DNA; DNA Damage; DNA Double Strand Break; DNA Repair; DNA Repair Gene; Defect; Detection; Disease; Event; Failure; Genetic Transcription; Genome; Genome Stability; Goals; Health; Histone H2A; Human; Individual; Ionizing radiation; Lesion; Link; Lysine; Mediating; Methionine; Modeling; Modification; Monoubiquitination; Mutation; N-terminal; Pathway interactions; Play; Polyubiquitin; Polyubiquitination; Post-Translational Protein Processing; Production; Proteins; Reader; Regulation; Role; Signal Transduction; Site; System; Transcript; Translations; Ubiquitin; Ubiquitination; ataxia telangiectasia mutated protein; cytotoxic; design; gene repression; genome integrity; improved; insight; novel; p53-binding protein 1; prevent; recruit; repaired; response; restriction enzyme; tool

ABSTRACT DNA double-strand breaks (DSBs) are one of the most deleterious lesions. The cellular responses to DSBs involve a sophisticated DNA damage response network that detects, signals and repairs the lesion; failure of this system results in mutation, deletion and other alteration of genome and underlies many diseases. Post- translational modifications by covalent attachment of ubiquitin to proteins, known as ubiquitination, play important regulatory roles in the DNA damage response. Previous studies, including ours, have illustrated that, upon DSBs detection and activation of the DNA damage response kinase ATM, Lys63-linked ubiquitination of histone H2A/H2AX on damaged chromatin is critical in recruiting DNA damage repair proteins, including 53BP1 and BRCA1, to the damage sites. However, much remains unknown about the complexity of ubiquitin modifications and their roles in the DNA damage response. We have discovered that Lys11-linkage–specific ubiquitin modification occurs on damaged chromatin that regulates repression of transcription at DSBs, revealing Lys11-linkage ubiquitin modification as a new signaling and regulatory platform in the response to DSBs. We further showed that this modification is ATM dependent and catalyzed by RNF8 and Ube2S. In addition, cells deficient in Lys11-linkage ubiquitin modification displayed increased sensitivity to ionizing radiation. It indicates a model that DNA repair and transcription can be regulated by distinct linkage-specific ubiquitin modifications at DSBs. However, it is not clear whether these two seemingly parallel linkage-specific ubiquitination pathways are coordinated to regulate DNA repair and transcription in the cellular responses to DSBs. The objective of this proposal is to identify the crosstalk between Lys63- and Lys11-linkage modification on damaged chromatin and determine the underlying mechanisms through which it coordinates transcriptional repression and DNA repair. We will pursue the following specific aims: 1) determine the crosstalk between Lys11- and Lys63-linkage ubiquitination mediated by Cezanne to regulate DNA repair; 2) determine additional mechanisms underlying the coordination of DNA repair and transcriptional inhibition; 3) determine the role of Lys11- and Lys63-linkage ubiquitination at defined DSB sites in regulating DNA repair and inhibition of transcription. Our findings should provide novel insights into the understanding of how ubiquitin modification at DSBs is involved in signaling events to regulate inhibition of transcription and repair. Our long-term goal is to decipher the complex DNA response network that protects genome integrity, which is essential for design of tools and treatment to improve human health.

抽象的 DNA 双链断裂 (DSB) 是最有害的损伤之一。细胞对 DSB 的反应 涉及复杂的 DNA 损伤反应网络，用于检测、发出信号并修复病变；的失败 该系统会导致基因组突变、缺失和其他改变，并成为许多疾病的根源。邮政- 通过泛素与蛋白质共价连接进行翻译修饰，称为泛素化，发挥作用 DNA 损伤反应中的重要调节作用。以前的研究，包括我们的研究，已经表明， DSB 检测并激活 DNA 损伤反应激酶 ATM 后，Lys63 泛素化 受损染色质上的组蛋白 H2A/H2AX 对于招募 DNA 损伤修复蛋白至关重要，包括 53BP1 和 BRCA1，到达损伤部位。然而，关于泛素的复杂性仍然有很多未知之处。 修饰及其在 DNA 损伤反应中的作用。我们发现 Lys11 连锁特异性 泛素修饰发生在受损染色质上，调节 DSB 转录抑制， 揭示 Lys11 连接泛素修饰作为响应的新信号和调节平台 DSB。我们进一步表明，这种修饰是 ATM 依赖性的，并由 RNF8 和 Ube2S 催化。在 此外，缺乏 Lys11 连接泛素修饰的细胞表现出对电离的敏感性增加 辐射。它表明DNA修复和转录可以通过不同的连锁特异性来调节的模型 DSB 处的泛素修饰。然而，尚不清楚这两个看似平行的联动是否特定 泛素化途径协调调节细胞响应中的 DNA 修复和转录 DSB。该提案的目的是确定 Lys63 和 Lys11 连锁修饰之间的串扰 受损染色质并确定其协调转录的潜在机制 抑制和 DNA 修复。我们将追求以下具体目标：1）确定之间的串扰 Cezanne 介导的 Lys11 和 Lys63 连锁泛素化调节 DNA 修复； 2) 确定附加 DNA 修复和转录抑制协调的机制； 3）确定角色 Lys11 和 Lys63 连锁泛素化在指定的 DSB 位点上调节 DNA 修复和抑制 转录。我们的研究结果应该为理解泛素修饰如何在 DSB 参与调节转录抑制和修复的信号事件。我们的长期目标是 破译保护基因组完整性的复杂 DNA 反应网络，这对于设计至关重要 改善人类健康的工具和治疗。