DNA–Protein Cross-Linking Sequencing for Genome-Wide Mapping of Abasic Sites at Single-Nucleotide Resolution

DNA-蛋白质交联测序，以单核苷酸分辨率进行全基因组脱碱基位点作图

• 批准号: 10723069
• 负责人: Feng Tang
• 金额: $ 11.91万
• 依托单位: UNIVERSITY OF CALIFORNIA RIVERSIDE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-08-11 至 2025-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10723069
• 关键词: Ablation; Acceleration; Affect; Age; Aging; Alkylation; Animal Model; Applications Grants; B-Lymphocytes; Bioinformatics; CRISPR/Cas technology; Cancer Etiology; Cells; Chromatin; Cytolysis; DNA; DNA Damage; DNA Repair; DNA glycosylase; DNA lesion; DNA-(apurinic or apyrimidinic site) lyase; DNA-Directed DNA Polymerase; DNA-protein crosslink; Data; Deamination; Depurination; Disease; Environmental Exposure; Enzymes; Epigenetic Process; Etiology; Failure; Generations; Genes; Genetic; Genome; Genome Mappings; Genomic DNA; Genomic Instability; Genomics; Glycols; Goals; Histones; Human; Human Genome; Hydrolysis; Knock-out; Knowledge; Learning; Malignant Neoplasms; Maps; Mediating; Mentors; Metabolism; Methods; Mutagenesis; Mutation; Mutation Analysis; Neurodegenerative Disorders; Nucleotides; Outcome; Pattern; Prevention strategy; Process; Publications; Publishing; Research; Research Project Grants; Resolution; Role; Scheme; Scientist; Site; Source; Testing; Therapeutic Intervention; Thymidine; Toxicant exposure; Toxicology; Training; Wild Type Mouse; Work; base; cancer genome; career; environmental toxicology; experience; genome sequencing; genome-wide; human disease; innovation; insight; mouse model; next generation sequencing; novel; oxidation; preventive intervention; repaired; skills; toxicant; whole genome

Project Summary Endogenous metabolism and environmental exposure can result in a battery of DNA lesions. Failure to repair these DNA lesions gives rise to genome instability and mutation accrual, which lead to accelerated aging, cancer, and neurodegenerative diseases. Abasic site (AP site) is one of the most prevalent forms of DNA damage. Although AP sites are known to be repaired by AP endonuclease in human cells, the dynamics of AP site induction and repair across the genome and the cellular factors modulating these processes remain elusive. In this application, we aim to explore how site-specific induction and repair of AP sites are affected by toxicant exposure and epigenetic alterations. We propose three specific aims to achieve our objective: 1) we will develop a DNA-protein cross-linking followed by next-generation sequencing (DPC-Seq) method for mapping, at single- base resolution, AP sites in the human genome after toxicant exposure; 2) we will explore how the formation and repair of AP sites are influenced by epigenetic state of chromatin; and 3) we will employ DPC-seq and whole- genome mutation analysis to examine how AP sites contribute to mutational signatures observed in cancer genomes. To accomplish this proposal and enable my transition to an independent academic career, I assembled a mentoring committee with expertise in animal models, bioinformatics, mutagenesis, and toxicology. The proposed research is built upon my research experience/skill sets, strong preliminary data, and the proposed mentoring plan. The outcome of the proposed research will establish a new method for mapping AP sites at single-base resolution and provide important insights into the occurrence and repair of AP sites and how they are influenced by toxicant exposure and genomic contexts. The proposed research will also reveal the roles of AP sites in cancer etiology, which will ultimately lead to better strategies for the prevention and therapeutic interventions of human cancer. Moreover, this proposal will fill gaps in my training and collect the data for my independent publications and research grant applications, thereby enabling me to transition to become an independent scientist in the field of environmental toxicology.

项目摘要 内源性代谢和环境暴露可导致一系列DNA损伤。未修理 这些DNA损伤引起基因组不稳定性和突变累积，这导致加速老化，癌症， 和神经退行性疾病。碱性位点（AP位点）是DNA损伤最普遍的形式之一。 虽然已知AP位点在人类细胞中被AP内切酶修复，但AP位点的动态变化不确定。 整个基因组的诱导和修复以及调节这些过程的细胞因子仍然难以捉摸。 在本申请中，我们的目的是探索如何特定的诱导和修复的AP网站的影响，毒物 暴露和表观遗传改变。我们提出了三个具体目标，以实现我们的目标：1）我们将发展 一种DNA-蛋白质交联，然后进行下一代测序（DPC-Seq）的作图方法， 基础分辨率，AP位点在人类基因组中的毒物暴露后; 2）我们将探讨如何形成和 AP位点的修复受染色质的表观遗传状态的影响;和3）我们将采用DPC-seq和全- 基因组突变分析，以检查AP位点如何有助于在癌症中观察到的突变特征 基因组为了完成这个建议，使我能够过渡到一个独立的学术生涯，我 组建了一个具有动物模型、生物信息学、诱变和毒理学专业知识的指导委员会。 拟议的研究是建立在我的研究经验/技能集，强大的初步数据，和拟议的 指导计划。 该研究成果将为单碱基AP位点定位提供一种新的方法 解决方案，并提供重要的见解，发生和修复的AP网站，以及他们如何受到影响， 和基因组背景的影响。拟议的研究还将揭示AP位点在 癌症病因学，这将最终导致更好的预防和治疗干预策略， 人类癌症此外，这项建议将填补我在培训中的空白，并为我的独立工作收集数据。 出版物和研究资助申请，从而使我能够过渡到成为一个独立的 环境毒理学领域的科学家。