Repair of DNA ends with adducts

用加合物修复 DNA 末端

• 批准号: 10587000
• 负责人: SANG EUN LEE
• 金额: $ 31万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-03-15 至 2027-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10587000
• 关键词: APEXL2 Gene; Address; Aging; Alleles; Antineoplastic Agents; BRCA deficient; BRCA1 gene; BRCA2 gene; Base Excision Repairs; Binding Proteins; Biochemical; Biochemical Genetics; Biochemistry; Biological; Biological Assay; Biological Models; Biological Process; Camptothecin; Cell Death; Cell physiology; Cells; Chemicals; Chromatin; Collaborations; Complex; DNA; DNA Adduction; DNA Adducts; DNA Damage; DNA Repair; DNA Repair Pathway; DNA lesion; DNA replication fork; DNA-(apurinic or apyrimidinic site) lyase; DNA-Directed DNA Polymerase; DNA-Directed RNA Polymerase; DNA-protein crosslink; Deoxyribonucleases; Development; Disease; Disease Resistance; Environmental Pollutants; Enzymes; Etiology; Excision; Exposure to; Formaldehyde; Foundations; Gene Expression; Genetic; Genetic Recombination; Genome; Genomic Instability; Genomics; Growth; Histones; Homologous Gene; Human; Interruption; Investigation; Laboratories; Lymphopoiesis; Maintenance; Malignant neoplasm of liver; Malignant neoplasm of ovary; Modeling; Molecular Genetics; Motor Neuron Disease; Movement; Mus; Mutagenesis; Mutation; Nerve Degeneration; Nucleotide Excision Repair; Nucleotides; Pathway interactions; Patients; Peptide Hydrolases; Peptides; Periodicity; Physiological; Process; Property; Proteins; Proteolysis; Reaction; Regulation; Repair Complex; Repetitive Sequence; Research; Resistance; Ribonucleotides; Role; Single Strand Break Repair; Stress; Symptoms; Tail; Technical Expertise; Testing; Therapeutic; Topoisomerase; Toxin; Type I DNA Topoisomerases; Tyrosine; Ubiquitin; Ubiquitination; Yeasts; adduct; anti-cancer; cancer cell; crosslink; environmental chemical; genetic approach; homologous recombination; human disease; innovation; inorganic phosphate; malignant breast neoplasm; multicatalytic endopeptidase complex; mutant; nuclease; prevent; protein complex; receptor; reconstitution; repaired; response; seal; therapy resistant

Project Summary Environmental pollutants and anti-cancer chemicals induce DNA–protein crosslinks (DPCs, also known as protein adducts) of topoisomerases and other proteins that obstruct almost every chromatin transaction and lead to mutations, genome instability and cell death if not removed quickly. Cells thus possess multiple mechanisms to remove DPCs: homologous recombination, nucleotide excision repair and proteolytic repair that involves proteolysis of bound proteins and nucleolytic cleavage of peptide-DNA bonds. Among these, proteolytic DPC repair is poorly defined and there are many questions about this unique collaboration between protease and nuclease and their substrate recognition. Using genetic and biochemical approaches, we have discovered that Apn2, the evolutionarily conserved back-up AP endonuclease, possesses an enzymatic activity unblocking DNA ends with various 3’ adducts including 2’,3’-cyclic phosphate, monophosphate and tyrosine DNA conjugates. We also found that Apn2 removes DNA-topoisomerase I crosslinks, the flagship DPC at 3' termini. The results might explain several puzzling symptoms in mice deleted for APE2, the metazoan Apn2 homolog, including dys-lymphopoiesis and growth retardation. The results also underscore the importance of 3’ blocked termini processing for genome maintenance and reveal unique challenges in removing these toxic DNA lesions in a tractable model system. Nevertheless, our results also raise a new set of questions about the regulation of Apn2 and APE2 in 3' adduct repair. For instance, we aim to determine how Apn2 recognizes a wide range of 3’ termini with different structural features and how such activities are regulated under unique cellular and functional contexts. We also want to elucidate the biological significance of Apn2 and human APE2-dependent Top1cc repair in cellular physology exposed to chemotherapeutics. Furthermore, we plan to define the roles of Apn2 and its interacting protease(s) in the removal of toxic DNA protein complexes (DPCs) that form via environmental pollutants and chemicals. The current proposal will address these important questions using innovative genetic and biochemical assays and shed light on the fundamental mechanisms of 3’ DNA adduct repair, mutagenesis and maintenance of chromosomal integrity. Emerging evidence suggests that accumulated DPCs cause aging, neurodegeneration and liver cancers. The human homolog of Apn2 is also essential for the viability of BRCA-deficient breast and ovarian cancer cells. The functional conservation in Top1cc and DPC repair between yeast and human further underscores the value of the proposed research in setting the foundation for analyzing the equivalent processes in human. Information gained from our proposed research plan will not only solve one of the fundamental questions in biological processes associated with mutagenesis, DNA repair, and genome maintenance, but will also impinge directly on the management and treatment of patients suffering from several devastating diseases with no clear treatment option.

项目摘要 环境污染物和抗癌化学品诱导DNA-蛋白质交联（DPC， 也称为蛋白加合物）的拓扑异构酶和其他蛋白质，阻碍几乎所有 染色质交易，并导致突变，基因组不稳定和细胞死亡，如果不删除 快因此，细胞具有多种去除DPC的机制：同源重组， 核苷酸切除修复和蛋白水解修复，涉及结合蛋白的蛋白水解， 肽-DNA键的核裂解。其中，蛋白水解DPC修复较差 关于蛋白酶和蛋白酶之间的这种独特的合作， 核酸酶及其底物识别。 利用遗传和生物化学方法，我们发现Apn 2，即进化上的 保守的备用AP核酸内切酶，具有解除封闭DNA末端的酶活性 与各种3'加合物，包括2'，3 '-环磷酸、单磷酸和酪氨酸DNA 结合物。我们还发现Apn 2去除了DNA-拓扑异构酶I交联， 在3'末端。这些结果可能解释了APE 2缺失小鼠的几个令人困惑的症状， 后生动物Apn 2同系物，包括淋巴细胞生成异常和生长迟缓。结果还 强调了3'封闭末端加工对基因组维持的重要性，并揭示了 在易处理的模型系统中去除这些毒性DNA损伤的独特挑战。 然而，我们的研究结果也提出了一系列关于Apn 2和APE 2调控的新问题。 在3'加合物修复中。例如，我们的目标是确定Apn 2如何识别广泛的3'， 具有不同结构特征的末端，以及这些活动如何在独特的 细胞和功能背景。我们还想阐明Apn 2的生物学意义， 暴露于化疗药物细胞生理学中的人类APE 2依赖性Top1 cc修复 此外，我们计划确定Apn 2及其相互作用的蛋白酶在去除 有毒的DNA蛋白复合物（DPC）是通过环境污染物和化学物质形成的。的 目前的建议将利用创新的遗传和生物化学技术来解决这些重要问题。 分析和阐明的基本机制，3' DNA加合物修复，诱变和 保持染色体的完整性。 新出现的证据表明，积累的DPC会导致衰老、神经退行性变和 肝癌Apn 2的人类同源物对于BRCA缺陷型细胞的生存能力也是必需的。 乳腺癌和卵巢癌细胞。Top1 cc和DPC修复中的功能保守性 酵母和人类进一步强调了拟议研究的价值， 用于分析人类的等效过程。从我们拟议的研究中获得的信息 该计划不仅将解决生物过程中的一个基本问题， 诱变，DNA修复和基因组维护，但也将直接影响 管理和治疗患有几种毁灭性疾病的患者， 治疗选择