Mechanisms of transcription regulation in chromatin

染色质转录调控机制

• 批准号: 9260911
• 负责人: JERRY L WORKMAN
• 金额: $ 41.25万
• 依托单位: STOWERS INSTITUTE FOR MEDICAL RESEARCH
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-05-01 至 2021-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9260911
• 关键词: ATP phosphohydrolase; Acetylation; Address; Affect; Binding Sites; Biochemistry; Cells; ChIP-seq; Chromatin; Chromatin Structure; Chromosome Structures; Code; Complex; Drosophila genus; Embryo; Gene Expression; Genes; Genetic; Genetic Transcription; Genomics; Goals; Histone Deacetylation; Histones; Human; Human Cell Line; Lead; Malignant Neoplasms; Mammalian Cell; Methyltransferase; Modification; Mutate; Mutation; Neurodegenerative Disorders; Nucleosomes; Open Reading Frames; Pathway interactions; Peptide Hydrolases; Proteins; Proteomics; RNA Polymerase II; Recurrence; SAGA; SCA7 protein; SWI/SNF Family Complex; Signal Transduction; Transcript; Transcription Coactivator; Transcriptional Regulation; Tumor Suppressor Proteins; Ubiquitin; Yeasts; dSWI/SNF; embryo tissue; fly; genome-wide; histone acetyltransferase; novel; public health relevance

 DESCRIPTION (provided by applicant): The goals of this proposal are to investigate fundamental mechanisms through which chromatin structure can regulate transcription and, when perturbed, creates conditions that lead to cancer. Studies will include the Swi/Snf and SAGA and complexes and the Set2/Rpd3S pathway in yeast, flies and human cells. The Swi/Snf nucleosome-remodeling complex functions as a tumor suppressor. Recurrent mutations in Swi/Snf are found in many cancers. To understand how loss of specific subunits affects Swi/Snf integrity and function, we will determine the Swi/Snf subunit interaction network in yeast and human cells. This information will uncover which subunits interact and form functional modules within the complex and how the functions of the complex change when subunits are missing or altered. We found that the Snf2 ATPase subunit of the yeast complex is modified by acetylation, and that this modification governs the dynamics of Swi/Snf occupancy genome wide. We will extend these studies to human Swi/Snf complexes to determine how they are regulated by acetylation. We will also purify Drosophila Swi/Snf complexes to determine its genomic binding sites and identify interacting proteins at different embryonic stages and in specific embryonic tissues. SAGA is a multisubunit histone acetyltransferase, histone ubiquitin protease and transcription co-activator. It contains several subunits with potential chromatin interacting domains and subunits that have been implicated in cancers and neurodegenerative disease. Major unresolved questions are which domains affect subunit functions and which localize SAGA to specific genes. We will address these questions by ChIP-Seq analysis after depleting specific subunits. We will explore novel proteins that interact with the ubiquitin protease module, which is no longer associated with SAGA in the absence of the Drosophila ataxin-7 subunit. We will pursue the function of this module and of intact SAGA in early zygotic transcription. The Set2 histone H3K36 methyltransferase, which functions as a tumor suppressor in human cells, is part of the Set2/Rpd3S pathway by which RNA polymerase II signals for histone deacetylation during transcription. We have found that acetylated histones accumulate in transcribed sequences because new acetylated histones replace the original histones in the absence of the Set2/Rpd3S pathway. Moreover, loss of Set2 leads to antisense transcription from within open reading frames. We will analyze the functions of Set2/Rpd3S pathway components in suppressing widespread antisense transcription. Conversely, we will analyze pathways by which antisense transcription represses coding gene transcripts, which also requires Set2, in mammalian cells. We will take both candidate and discovery approaches to identify novel Set2 targets. In addition, we will analyze the effects of cancer-associated mutations in Set2 with Set2 function in yeast and human cell lines. 1

 描述（由适用提供）：该提案的目标是研究染色质结构可以调节转录的基本机制，并且在扰动时会产生导致癌症的疾病。研究将包括SWI/SNF和SAGA以及复合物以及酵母，苍蝇和人类细胞中的SET2/RPD3S途径。 SWI/SNF核能复合物复合物作为肿瘤抑制剂。在许多癌症中发现了SWI/SNF中的复发突变。为了了解特定亚基的损失如何影响SWI/SNF的完整性和功能，我们将确定酵母和人类细胞中的SWI/SNF亚基相互作用网络。该信息将发现哪些亚基在复合物中相互作用并形成功能模块，以及在丢失或更改亚基时复合物的功能如何变化。我们发现，酵母复合物的SNF2 ATPase亚基是通过乙酰化修饰的，并且这种修饰控制了SWI/SNF占用基因组宽的动力学。我们将将这些研究扩展到人类SWI/SNF复合物，以确定如何通过乙酰化调节它们。我们还将纯化果蝇SWI/SNF复合物来确定其基因组结合位点，并在不同的胚胎阶段和特定的胚胎组织中鉴定相互作用的蛋白质。传奇是一种多亚基组蛋白乙酰转移酶，组蛋白泛素蛋白和转录共激活剂。它包含几个具有潜在染色质相互作用结构域的亚基和癌症和神经退行性疾病中隐含的亚基。主要未解决的问题是哪些领域影响亚基功能，哪些域将传奇定位于特定基因。在耗尽特定亚基后，我们将通过CHIP-seq分析来解决这些问题。我们将探索与泛素蛋白酶模块相互作用的新型蛋白质，该蛋白质酶模块不再与果蝇亚Xaxin-7亚基相关联。我们将在早期的合子转录中追求该模块和完整传奇的功能。 SET2组蛋白H3K36甲基转移酶在人类细胞中充当肿瘤抑制剂，是SET2/RPD3S途径的一部分，在转录过程中，RNA聚合酶II信号用于组蛋白脱乙酰基化。我们发现，乙酰化组蛋白会积聚在转录序列中，因为在没有SET2/RPD3S途径的情况下，新的乙酰化组蛋白代替了原始组蛋白。此外，SET2的丢失会导致从开放式阅读框内进行反义转录。我们将分析SET2/RPD3S途径成分的功能，以抑制宽度的反义转录。相反，我们将分析反义转录反映哺乳动物细胞中需要SET2的编码基因转录物的途径。我们将采用候选人和发现方法来识别新颖的SET2目标。此外，我们将分析与酵母和人类细胞系中SET2功能的SET2相关突变的影响。 1