Administrative supplement to support investigation into the structural basis of ubiquitin signaling in response to DNA alkylation damage

支持调查泛素信号传导响应 DNA 烷基化损伤的结构基础的行政补充

• 批准号: 10580459
• 负责人: Patrick Lombardi
• 金额: $ 4.96万
• 依托单位: MOUNT ST. MARY'S UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-15 至 2023-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10580459
• 关键词: Address; Administrative Supplement; Affect; Affinity; Alkylating Agents; Alkylation; Binding; Binding Sites; Biochemistry; Biophysics; Calorimetry; Cell Death; Cell physiology; Cells; Cellular biology; Chemistry; Collaborations; Color; Complex; DNA Alkylation; DNA Damage; DNA Modification Process; DNA Repair; DNA Repair Pathway; DNA Replication Inhibition; DNA Sequence Alteration; DNA lesion; Defect; Dependence; Detection; Diagnosis; Disease; Event; Exposure to; First Generation College Students; Genomic Instability; Goals; Grant; Human body; Investigation; Knock-out; Knowledge; Left; Link; Lysine; Measures; Modification; Molecular; Mutate; NMR Spectroscopy; Pathway interactions; Polyubiquitin; Population Biology; Principal Investigator; Process; Proliferating; Property; Proteins; Protomer; Repair Complex; Research; Research Personnel; Research Proposals; Side; Signal Transduction; Site; Source; Specificity; Structure; Students; Therapeutic; Time; Titrations; UBD protein; Ubiquitin; Universities; X-Ray Crystallography; biophysical properties; biophysical techniques; cancer therapy; chemotherapy; cytotoxic; experimental study; lower income families; mutant; novel; nucleobase; prevent; recruit; repair enzyme; repaired; response; structural biology; three dimensional structure; tumor; undergraduate student

Project Summary/Abstract DNA alkylation damage comprises a class of prevalent, harmful nucleobase modifications that occur thousands of times per cell per day in the human body as a result of endogenous and exogenous sources. Left unrepaired, DNA alkylation damage can result in genetic mutations, the inhibition of DNA replication, and cell death. Several DNA repair pathways have evolved to reverse the numerous DNA modifications that result from alkylation damage. While the repair enzymes in these pathways are well studied, much less is known about the upstream signaling events that initiate DNA repair and localize repair complexes to damage sites. It was recently shown that the ALKBH3-ASCC DNA repair complex is recruited to alkylation damage sites by binding chains of the protein ubiquitin that are assembled in proximity to the DNA lesions. The protein ASCC2 is responsible for binding the polyubiquitin chains that localize the ALKBH3-ASCC complex. A vast array of different types of polyubiquitin chains are present in cells, however, and it is unclear how ASCC2 selectively recognizes the K63- linked polyubiquitin chains that signal alkylation damage. The PI proposes to use a combination of structural biology, cell biology, and biophysics to investigate ASCC2’s selectivity for K63-linked polyubiquitin chains and the dependence of ALKBH3-ASCC complex localization on the unique ubiquitin-binding properties of ASCC2. The specific aims of the project are: 1) to identify the novel ASCC2:ubiquitin binding interface that imparts enhanced affinity for polyubiquitin chains, 2) to determine the structural basis of ASCC2’s specificity for binding K63-linked polyubiquitin chains, and 3) to quantify the contribution of ASCC2’s ubiquitin-binding properties to DNA alkylation damage repair. Investigating the outstanding questions associated with DNA alkylation damage repair will allow clinicians to better understand diseases that result from defects in alkylation damage repair pathways and to more effectively deploy alkylating agents as therapeutics, especially for the treatment of cancer. Furthermore, these experiments will also provide valuable research opportunities for students at Mount St. Mary’s University (MSMU), where substantial populations of the biology, chemistry, and biochemistry majors are first-generation college students (16.7%), students of color (42.5%), or students from moderate- or low-income families (27.1 % Pell Grant recipients). Overall, the proposed experiments will address a lack of knowledge in the current understanding of DNA alkylation damage repair while greatly enhancing research opportunities for students at MSMU.

项目摘要/摘要 DNA烷基化损伤包括一类普遍的有害核碱酶修饰 由于内源性和外源性来源，人体每个细胞的每日时间都有时间。剩下的 DNA烷基化损伤会导致基因突变，DNA复制的抑制和细胞死亡。一些 DNA修复途径已演变为逆转毒品导致的大量DNA修饰 损害。虽然这些途径中的维修酶是很好的研究，但对上游知之甚少 启动DNA修复并将修复复合物定位到损坏位点的信号事件。最近显示了 ALKBH3-ASCC DNA修复复合物是通过结合链的 与DNA病变接近的蛋白泛素。蛋白质ASCC2负责 结合定位ALKBH3-ASCC复合物的多泛素链。一系列不同类型的 但是，多泛素链存在于细胞中，尚不清楚ASCC2如何选择地识别K63- 链接的多泛素链，这些链信号烷基化损伤。 PI提议结合结构的组合 生物学，细胞生物学和生物物理学，以研究ASCC2对K63连接的多泛素链的选择性和 ALKBH3-ASCC复合物定位对ASCC2独特的泛素结合特性的依赖性。 该项目的具体目的是：1）识别新颖的ASCC2：泛素结合界面 增强对多泛素链的亲和力，2）确定ASCC2结合特异性的结构基础 K63连接的多泛素链，以及3）量化ASCC2的泛素结合特性对 DNA烷基化损伤修复。调查与DNA烷基化损伤相关的未取得的问题 维修将使临床医生能够更好地了解酗酒损害修复缺陷导致的疾病 途径，更有效地将烷基化剂作为治疗，特别是用于治疗癌症。 此外，这些实验还将为Mount St.的学生提供宝贵的研究机会。 玛丽大学（MSMU），生物学，化学和生物化学专业的大量人口是 第一代大学生（16.7％），有色学生（42.5％）或中等或低收入的学生 家庭（27.1％的佩尔赠款接收者）。总体而言，拟议的实验将解决缺乏 当前对DNA烷基化损伤修复的理解，同时大大增加了研究机会 MSMU的学生。