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Epigenetic mechanisms of Ash1L in transcriptional activation

Ash1L 在转录激活中的表观遗传机制

基本信息

批准号：
10399431
负责人：
Jin He
金额：
$ 33.36万
依托单位：
MICHIGAN STATE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-05-01 至 2025-04-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10399431
关键词：
ASH1L gene Address Binding Biochemical Biological Assay CRISPR/Cas technology Cells Complex Cryoelectron Microscopy Development Disease Drosophila genus ES Cell Line Enzymes Epigenetic Process Event Gene Activation Gene Expression Genes Genetic Genetic Transcription Genetic study Histone H3 Histones Homologous Gene Impairment In Vitro Individual Lead Link Lysine MLL gene Malignant Neoplasms Mammalian Cell Mammals Mediating Methylation Mixed-Lineage Leukemia Modeling Modification Molecular Mus Mutate Nucleic Acid Regulatory Sequences Phenotype Play Polycomb Process Proteins Reader Research Institute Role Structure Tail Time Transcriptional Activation base design embryonic stem cell epigenetic regulation gene repression genetic analysis genome editing histone methylation histone methyltransferase histone modification inhibitor mutant new therapeutic target next generation sequencing promoter recruit stem cell differentiation transcription factor

项目摘要

Project Abstract Epigenetic modifications at transcriptional regulatory regions play an important role in facilitating lineage-specific gene expression during stem cell differentiation. In addition to lineage-specific transcription factors, Trithroax-group (TxG)-group proteins promote lineage-specific gene expression through antagonizing the Polycomb-mediated transcriptional repression. In mammals, Mll1/Mll2 (Mixed Lineage Leukemia) and Ash1L (Absent, Small, or Homeotic discs 1-Like) complexes are two TxG complexes that mediate covalent histone modifications through their histone methyltransferase (HMTase) activities towards histone H3 lysine 4 and histone H3 lysine 36 respectively. Although previous genetic studies have revealed that Mll1/Mll2 and Ash1L complexes are functionally involved in a common epigenetic regulatory process, it remains unknown how these two complexes are connected at the molecular level to carry out their functions in transcriptional activation. Additionally, in contrast to the well-studied Mll1/Mll2 complexes and histone H3K4 methylation, the functions of Ash1L and its mediated histone H3K36 methylation at promoters are largely unknown. In an effort to address these fundamental questions, we purified the Ash1L-interacting proteins and identified Spindlin1 (Spin1), a histone H3K4me3-specific reader, physically binds to Ash1L. Deletion of either Ash1L or Spin1 in mouse embryonic stem cells impairs the expression of early lineage-specific genes upon induced differentiation, suggesting a functional connection between Spin1 and Ash1L in cells. Built upon these results, we propose a new model to unify the function of Mll1/Mll2 and Ash1L in transcriptional activation. In this model, Mll1/Mll2, Spin1, and Ash1L form an epigenetic regulatory axis, in which the individual component is sequentially recruited to the lineage-specific gene promoters to mediate histone modifications during transcriptional activation. Specifically, Spin1 plays a central role in connecting Mll1/Mll2 and Ash1L by recruiting Ash1L to the H3K4me3-marked lineage-specific gene promoters. To understand the functional role of individual components in this Mll1/Mll2-Spin1-Ash1L epigenetic regulatory axis, we will combine biochemical assays, CRISPR/Cas9- mediated genome editing, and next generation sequencing-based genetic analysis to dissect individual regulatory step proposed in the working model. Specifically, we will determine (1) whether Ash1L and its HMTase activity are required for its function in facilitating transcriptional activation; (2) the molecular mechanisms for the recruitment of Ash1L to gene promoters; (3) the structural basis underlying the interaction between Ash1L, Spin1 and histone H3K4me3. Completion of this study will not only significantly advance our understanding on the basic epigenetic mechanisms regulating the lineage-specific gene activation, but also reveal new therapeutic targets for blocking aberrant gene activation in cancers.

项目摘要转录调控区的表观遗传修饰在促进谱系特异性表达中起重要作用。干细胞分化过程中的基因表达。除了谱系特异性转录因子， Trithroax-group（TxG）-group蛋白通过拮抗多梳介导的转录抑制。在哺乳动物中，Mll 1/Mll 2（混合谱系白血病）和Ash 1 L （缺失、小或同源异型盘1样）复合物是两种TxG复合物，其介导共价组蛋白通过其对组蛋白H3赖氨酸4的组蛋白甲基转移酶（HMTase）活性进行修饰，组蛋白H3赖氨酸36。尽管先前的遗传学研究已经揭示了Mll 1/Mll 2和Ash 1 L 复合物在功能上参与了一个共同的表观遗传调控过程，目前还不清楚这些复合物是如何被激活的。两个复合物在分子水平上连接，以实现它们在转录激活中的功能。此外，与研究充分的Mll 1/Mll 2复合物和组蛋白H3 K4甲基化相反， Ash 1 L及其介导的组蛋白H3 K36启动子甲基化在很大程度上未知。为了解决为了解决这些基本问题，我们纯化了Ash 1 L相互作用蛋白，并鉴定了Spindlin 1（Spin 1），组蛋白H3 K4 me 3特异性阅读器，与Ash 1 L物理结合。小鼠中Ash 1 L或Spin 1的缺失胚胎干细胞在诱导分化时损害早期谱系特异性基因的表达，这表明细胞中Spin 1和Ash 1 L之间存在功能联系。基于这些结果，我们提出了一个新的模型来统一Mll 1/Mll 2和Ash 1 L在转录激活中的功能。在该模型中，Mll 1/Mll 2， Spin 1和Ash 1 L形成了一个表观遗传调控轴，其中单个组分被顺序募集谱系特异性基因启动子介导转录激活过程中的组蛋白修饰。具体而言，Spin 1通过将Ash 1 L募集到Mll 1/Mll 2和Ash 1 L中而在连接Mll 1/Mll 2和Ash 1 L中起核心作用。 H3 K4 me 3标记的谱系特异性基因启动子。了解各个组件的功能作用在这个Mll 1/Mll 2-Spin 1-Ash 1 L表观遗传调控轴中，我们将联合收割机生化测定、CRISPR/Cas9- 介导的基因组编辑和下一代基于测序的遗传分析，工作模式中提出的监管步骤。具体而言，我们将确定（1）Ash 1 L及其 HMTase活性是其促进转录激活的功能所必需的;（2）HMTase的分子生物学活性， Ash 1 L募集到基因启动子的机制;（3）相互作用的结构基础 Spin 1和组蛋白H3 K4 me 3之间的关系。这项研究的完成不仅将大大提高我们的了解调节谱系特异性基因激活的基本表观遗传机制，揭示了阻断癌症中异常基因激活的新治疗靶点。