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会导致一系列DNA病变。无法修复 这些DNA病变会导致基因组不稳定性和突变应计，从而导致衰老，癌症，， 和神经退行性疾病。 Abasic位点（AP位点）是DNA损伤最普遍的形式之一。 尽管已知AP位点通过人类细胞中的AP核酸内切酶修复，但AP位点的动力学 整个基因组和调节这些过程的细胞因子的诱导和修复仍然难以捉摸。 在此应用中，我们旨在探讨AP站点的特定地点诱导和维修如何受到毒物的影响 暴露和表观遗传改变。我们提出了三个特定目标以实现我们的目标：1）我们将发展 DNA-蛋白质交联，然后是下一代测序（DPC-SEQ）用于映射的方法 基础分辨率，毒性暴露后人类基因组中的AP位点； 2）我们将探讨形成和 AP位点的修复受染色质表观遗传状态的影响； 3）我们将采用dpc-seq和整个 基因组突变分析以检查AP位点如何促进癌症观察到的突变特征 基因组。为了实现这一建议并使我过渡到独立的学术生涯，我 组建了一个指导委员会，该委员会在动物模型，生物信息学，诱变和毒理学方面具有专业知识。 拟议的研究建立在我的研究经验/技能，强大的初步数据以及拟议的 指导计划。 拟议的研究的结果将建立一种新方法，用于映射单基础的AP位点 解决方案并提供有关AP站点的发生和维修及其如何影响的重要见解 通过毒物暴露和基因组环境。拟议的研究还将揭示AP站点在 癌症病因，最终将为预防和治疗干预措施提供更好的策略 人类癌。此外，该建议将填补我的培训空白，并为我的独立 出版和研究赠款申请，从而使我能够过渡成为独立的 环境毒理学领域的科学家。