Control mechanisms of 5-hydroxymethylcytosine metabolism in human cells

人体细胞5-羟甲基胞嘧啶代谢的控制机制

• 批准号: 10629908
• 负责人: Haeyoung Kim
• 金额: $ 13.8万
• 依托单位: TEXAS A&M UNIVERSITY-KINGSVILLE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-01 至 2027-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10629908
• 关键词: Attention; Base Excision Repairs; Biological Assay; Biomedical Research; Blood; Brain; CRISPR/Cas technology; Cell Line; Cell physiology; Cells; CpG dinucleotide; Cytosine; DNA; DNA Damage; DNA Double Strand Break; DNA Methylation; DNA Modification Methylases; DNA Repair; DNA biosynthesis; DNA lesion; Development; Disease; Enzymes; Epigenetic Process; Family; Functional disorder; Gene Expression Regulation; Generations; Genes; Genetic Transcription; Genome; Genome Stability; Glioblastoma; Goals; Human; Induced pluripotent stem cell derived neurons; Knowledge; Light; Link; Lymphoma; Malignant Neoplasms; Mass Spectrum Analysis; Measures; Mediating; Metabolism; Methylation; Mitotic; Modeling; Neurodegenerative Disorders; Neurons; Nuclear; Nucleotides; Organ; Organism; Outcome; Pathway interactions; Pluripotent Stem Cells; Process; Production; Proteins; Resistance; Role; Site; Survival Rate; System; Techniques; Testing; Thymine DNA Glycosylase; Time; Tissue-Specific Gene Expression; Tissues; age related; anti-cancer therapeutic; cell type; conditional knockout; demethylation; drug discovery; epigenetic marker; epigenetic regulation; high throughput screening; human DNA; human disease; knock-down; mortality; novel; novel marker; novel therapeutics; oxidation; postmitotic; repaired; response; translocase; tumor progression

Project Summary The 5-hydroxymethylcytosine (5hmC) in mammalian DNA is drawing significant attention in epigenetics because of its indispensable roles in gene expression regulation. The loss of 5hmCs in various cancers including glioblastomas and lymphomas has been linked to the poor survival rate as well as the resistance to anti-cancer therapeutics. Although there is amassing evidence of 5hmCs as a novel epigenetic marker, there is very little understanding on how 5hmCs are enriched at specific loci in the genome. In this project, we aim to identify the factors controlling the DNA demethylation pathway, which result in the formation of locus-specific 5hmCs in human neurons. For this purpose, proteins controlling the activity of ten-eleven translocase (TET) family enzymes in the DNA demethylation pathway will be identified. To examine the function of candidate proteins, we will take advantage of human induced-neurons (hiNs) derived from human induced-pluripotent stem cells (hiPSCs). In addition, we will elucidate the role and fate of 5hmCs formed during DNA damage repair process using blue light-inducible CRISPR/Cas9 technique. This robust experimental platform will enable the rapid production of isogenic and homogenous neurons and other cell types. With this cell-based assay system, high-throughput assays can be developed that will advance biomedical researches and drug discoveries. Finally, we expect the successful completion of this project will expand our understanding on DNA demethylation and their roles in epigenetics, development, and various human diseases.

项目摘要 哺乳动物DNA中的5-羟甲基胞嘧啶（5-hydroxymethylcytosine，5 hmC）是表观遗传学研究的热点 因为它在基因表达调控中起着不可或缺的作用。各种癌症中5 hmCs的丢失 包括胶质母细胞瘤和淋巴瘤与低存活率以及对 抗癌疗法。尽管有大量证据表明5 hmCs是一种新的表观遗传标记， 对5 hmC如何在基因组中的特定位点富集的了解很少。在这个项目中，我们的目标是 确定控制DNA去甲基化途径的因素，这些因素导致形成基因座特异性 人神经元中的5 hmCs。为此目的，控制10 - 11易位酶（泰特）活性的蛋白质 将鉴定DNA去甲基化途径中的家族酶。考察候选人的功能 蛋白质，我们将利用人类诱导神经元（hiN）来源于人类诱导多能 干细胞（hiPSC）。此外，我们将阐明在DNA损伤过程中形成的5 hmCs的作用和命运 使用蓝光诱导的CRISPR/Cas9技术的修复过程。这个强大的实验平台将 能够快速产生同基因和同质的神经元和其他细胞类型。有了这种基于细胞的 分析系统，可以开发高通量分析，这将促进生物医学研究和药物 发现。最后，我们期望这个项目的成功完成将扩大我们对DNA的理解 去甲基化及其在表观遗传学、发育和各种人类疾病中的作用。