Role of NEIL DNA glycosylases in DNA demethylation and differentiation

NEIL DNA 糖基化酶在 DNA 去甲基化和分化中的作用

• 批准号: 319444698
• 负责人: Professor Dr. Christof Niehrs, Ph.D.
• 金额: --
• 依托单位: Abteilung Molekulare Embryologie
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2016
• 资助国家: 德国
• 起止时间: 2015-12-31 至 2019-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/319444698?language=en
• 关键词: Role; NEIL; DNA; glycosylases; demethylation

DNA methylation is an epigenetic mark which plays important roles in development and disease. It is now widely accepted that DNA methylation is dynamic and is subject to enzymatic demethylation. In the TET/TDG demethylation pathway, methylated cytosine is iteratively oxidized by TET dioxygenases and unmodified cytosine is restored by thymine DNA glycosylase (TDG). A vulnerable intermediate in TET/TDG demethylation is the abasic (AP) site. We have shown recently that NEIL1 and NEIL2 DNA glycosylases coordinate AP site processing during TET/TDG DNA demethylation. Our analysis indicates that NEILs function as regulators in the coordinated substrate hand-over during oxidative DNA demethylation. To investigate the physiological relevance and biological role of Neil DNA glycosylases in DNA demethylation and DNA repair we have generated via CRISPR/Cas9, mouse embryonic stem cells which are mutant for Neil1,-2, or 3, as well as Neil double mutants The aim of this project is to characterize these mutant ES cells for their differentiation capacity, oxidized cytosine levels, and DNA repair abilities.

DNA甲基化是一种表观遗传标记，在发育和疾病中起重要作用。现在普遍认为DNA甲基化是动态的，并且受到酶促去甲基化的影响。在泰特/TDG去甲基化途径中，甲基化的胞嘧啶被泰特双加氧酶反复氧化，未修饰的胞嘧啶被胸腺嘧啶DNA糖基化酶（TDG）恢复。泰特/TDG去甲基化中的易受攻击的中间体是脱碱基（AP）位点。我们最近发现NEIL 1和NEIL 2 DNA糖基化酶在泰特/TDG DNA去甲基化过程中协调AP位点加工。我们的分析表明，NEIL的功能作为调节剂在协调的底物移交过程中的氧化DNA去甲基化。为了研究Neil DNA糖基化酶在DNA去甲基化和DNA修复中的生理相关性和生物学作用，我们通过CRISPR/Cas9产生了Neil 1，2或3突变的小鼠胚胎干细胞，以及Neil双突变体。本项目的目的是表征这些突变ES细胞的分化能力，氧化胞嘧啶水平和DNA修复能力。