Role of BAP1/ASXL3 complex in transcriptional regulation and development

BAP1/ASXL3 复合物在转录调控和发育中的作用

• 批准号: 10669750
• 负责人: Lu Wang
• 金额: $ 40万
• 依托单位: NORTHWESTERN UNIVERSITY AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-07-22 至 2027-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10669750
• 关键词: Attenuated; Biochemical; Biological Markers; Bromodomain; Cells; Chromatin; Chromatin Structure; Complex; Development; Developmental Process; Disease; Enhancers; Epigenetic Process; Equilibrium; Functional disorder; Gene Expression; Genes; Genetic; Genetic Transcription; Goals; Histone H2A; Histones; Lysine; Malignant Neoplasms; Mammalian Cell; Mammals; Mediating; Modification; Molecular; Multiprotein Complexes; Mus; Mutation; Neurons; PRC1 Protein; Patients; Play; Polycomb; Prognostic Marker; Proteins; Role; Scaffolding Protein; Therapeutic; Transcription Coactivator; Transcriptional Regulation; Ubiquitination; developmental disease; driver mutation; embryonic stem cell; gene repression; histone modification; human disease; nerve stem cell; new therapeutic target; novel; novel therapeutics; recruit; screening; small molecule; small molecule inhibitor; stem cell differentiation; transcriptome; ubiquitin isopeptidase

Project Summary Recent advances have shown that dysfunctions prevalent in epigenetic factors play an important role in the developing human diseases, including developmental disorders and cancers. Therefore, understanding the roles of these epigenetic factors may aid in identifying new epigenetic prognostic markers or targetable biomarkers that could contribute towards the establishment of novel therapeutics. Histone H2A lysine 119 ubiquitination (H2AK119ub) is one of the most functional histone marks that plays an essential role in establishing repressive chromatin domains and mediating Polycomb induced transcriptional repression during development. The evolutionarily conserved H2AK119ub histone modification is mainly catalyzed by the Polycomb repressive complex 1 (PRC1) and deubiquitinated by Polycomb repressive-deubiquitinase (PR-DUB) complex (also known as the BAP1 complex in mammals) that antagonizes PRC1’s function. In mammalian cells, the BAP1 complex functions as a multi-protein complex, containing as many as ten different subunits, which are responsible for the chromatin recruitment, protein stability, and enzymatic activity of BAP1. Mutations and dysregulations within subunits in the BAP1 complex are found in patients with developmental diseases, neuronal disorders, and cancer. Therefore, it is critical to understand the molecular basis of how genes are turned on/off by the BAP1/PRC1 epigenetic balance. Our previous studies have characterized a functional epigenetic axis comprised of both BAP1 and Bromodomain and Extraterminal (BET)-containing protein 4 (BRD4), which are physically bridged together by the scaffold protein ASXL3—the largest subunit within the BAP1 complex. Genetic depletion of the linker, ASXL3, dramatically attenuates the establishment of the BAP1/ASXL3/BRD4 epigenetic axis machinery at active enhancers, leading towards a significant reduction in the enhancer-nearby gene expression. In our current studies, we have employed biochemical, molecular, and small-molecule screening approaches to mechanistically understand how the ASXL3/BAP1/BRD4 epigenetic axis regulates transcription and determines cell fate and differentiation ability. Our first goal for this study is to uncover the BAP1/ASXL3 sub-complex’s role at active enhancers and assess the impact of the BAP1/ASXL3/BRD4 epigenetic axis on enhancer activity, chromatin structure, and gene expression. As a major H2AK119 deubiquitinase, the BAP1 complex functions as a general transcriptional activator, antagonizing PRC1’s function and is involved in PRC1-dependent transcriptional regulation. Therefore, our second project is to elucidate the relationship between BAP1 and a distinct PRC1 sub-complex in regulating H2AK119ub levels by utilizing our newly developed BAP1-specific small-molecule inhibitor. Mutations within ASXL3 have been demonstrated to be driver mutations in multiple neuronal diseases. Therefore, we will define the role of ASXL3 in mediating ESC differentiation into neuronal progenitor cells (NPC), and then investigate how ASXL3 and its associated epigenetic factors determine the transcriptome landscape during developmental processes.

项目摘要 最近的研究表明，表观遗传因子中普遍存在的功能障碍在 发展人类疾病，包括发育障碍和癌症。因此，理解这些角色 这些表观遗传因素可能有助于识别新的表观遗传预后标志物或靶向生物标志物 这可能有助于建立新的疗法。组蛋白H_2A赖氨酸119泛素化 (H2AK119ub)是功能最强的组蛋白标志物之一，在建立抑制机制中起着至关重要的作用 染色质结构域和中介多梳在发育过程中诱导转录抑制。这个 进化上保守的组蛋白修饰主要是由多梳抑制基因催化的 由多梳抑制-脱泛素酶(PR-DUB)复合体(也称为 如哺乳动物中的BAP1复合体)，拮抗Prc1‘S的功能。在哺乳动物细胞中，BAP1复合体 作为一个多蛋白质复合体，包含多达10个不同的亚基，负责 染色质募集、蛋白质稳定性和BAP1的酶活性。体内的突变和失调 BAP1复合体中的亚单位在发育疾病、神经元障碍和 癌症。因此，了解基因如何由基因启动/关闭的分子基础至关重要。 BAP1/Prc1表观遗传平衡。我们之前的研究已经表征了一个功能性表观遗传轴，包括 BAP1和溴结构域以及含有BET的蛋白4(BRD4)，它们都是物理上 通过支架蛋白ASXL3-BAP1复合体中最大的亚基-连接在一起。基因枯竭 连接子ASXL3的存在显著减弱了BAP1/ASXL3/BRD4表观遗传轴的建立 作用于活性增强子的机制，导致增强子附近的基因表达显著减少。 在我们目前的研究中，我们使用了生化、分子和小分子筛选方法来 从机制上理解ASXL3/BAP1/BRD4表观遗传轴如何调节转录并决定 细胞命运和分化能力。我们这项研究的第一个目标是揭示BAP1/ASXL3亚复合体的作用 在活性增强子并评估BAP1/ASXL3/BRD4表观遗传轴对增强子活性的影响， 染色质结构和基因表达。作为一种主要的H_2AK119脱泛素酶，BAP1复合体的功能如下 一种通用转录激活因子，拮抗Prc1‘S功能，参与Prc1依赖 转录调控。因此，我们的第二个项目是阐明BAP1和a之间的关系 利用我们新开发的BAP1特异性蛋白调节H_2AK119ub水平的不同PRC1亚复合体 小分子抑制剂。ASXL3内的突变已被证明是多发性硬化症的驱动因素 神经性疾病。因此，我们将确定ASXL3在介导胚胎干细胞分化为神经元中的作用 ，然后研究ASXL3及其相关的表观遗传因素如何决定 发育过程中的转录组景观。