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.

项目概要 哺乳动物 DNA 中的 5-羟甲基胞嘧啶 (5hmC) 引起了表观遗传学的极大关注 因为它在基因表达调控中发挥着不可或缺的作用。各种癌症中 5hmC 的丢失 包括胶质母细胞瘤和淋巴瘤在内的癌症与低生存率以及耐药性有关 抗癌疗法。尽管有大量证据表明 5hmCs 作为一种新型表观遗传标记，但 对于 5hmC 如何在基因组中的特定位点富集，我们知之甚少。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，从而导致位点特异性的形成 人类神经元中的 5hmC。为此，控制十一十一转位酶 (TET) 活性的蛋白质 DNA 去甲基化途径中的家族酶将被鉴定。考察候选人的职能 蛋白质，我们将利用源自人类诱导多能神经元（hiN）的优势 干细胞（hiPSC）。此外，我们将阐明 DNA 损伤过程中形成的 5hmC 的作用和命运 使用蓝光诱导 CRISPR/Cas9 技术进行修复过程。这个强大的实验平台将 能够快速产生等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。