Structural and functional determinants of biomolecular condensates in transcription organization

转录组织中生物分子凝聚体的结构和功能决定因素

• 批准号: 10714536
• 负责人: Jan-Hendrik Spille
• 金额: $ 33.92万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-22 至 2028-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10714536
• 关键词: Binding Sites; Biological Process; Cell Nucleus; Cell physiology; Cells; Chromatin; Chromatin Structure; Data; Development; Disease; Elements; Enhancers; Epigenetic Process; Genetic Transcription; Genomics; Health; Informal Social Control; Knowledge; Link; Membrane; Microscopy; Modeling; Modification; Molecular; Mus; Nature; Nuclear; Organelles; Phase; Physical condensation; Proteins; Regulatory Element; Research; Resolution; Structure; Surface; Testing; Time; Transcriptional Regulation; Vision; biophysical model; embryonic stem cell; genome-wide analysis; programs; scaffold; self organization; superresolution microscopy; transcription factor

Many cellular processes use phase separation to sequesters biomolecules into membraneless organelles or condensates. This concept is of particular importance in the cell nucleus. Most nuclear condensates are associated with specific chromatin loci. Thus, chromatin organization and biomolecular condensate formation are closely related. Topology, epigenetic modification, or enzymatic activity are possible links between the chromatin scaffold and protein condensation. We have previously described condensates that concentrate transcription machinery at super-enhancers but understanding of cell-to-cell variability and locus specific features that nucleate condensates is missing. Preliminary data indicates that active chromatin decorates the surface of transcription condensates below the resolution limit of conventional microscopy. Making use of recent developments in multiplexed super-resolution microscopy of both chromatin elements and protein factors, we will characterize the nature of the chromatin enrichment layer and determine if it reflects the condensate surface as an active interface that regulates transcription. We will for the first time directly observe the interplay of local chromatin topology, epigenetic signatures, and condensate formation in a specific model locus in mouse embryonic stem cells. By scrutinizing with genomic resolution how each of the regulatory elements in the locus interacts with the condensate we will determine if chromatin structures such as enhancer hubs nucleate condensates, or if condensates induce specific chromatin topologies by connecting active elements. Finally, we will investigate at the molecular level how condensate constituents interact with the microenvironment and associated chromatin elements, and test the hypothesis that retention of transcription factors boosts binding site occupancy and stabilizes condensates at regulatory chromatin. Targeted perturbation and locus-specific observation will allow us to answer this question and begin to identify genome-wide rules that predict where precisely chromatin-supported condensates form. The overall vision for this research program is to develop an integrated view of chromatin organization and condensate formation. Experimental data will inspire new conceptual frameworks and help us integrate the full complexity of condensates into paradigms of biological function such as transcription regulation. We expect that new biophysical models of self-organization and self-regulation will emerge from our studies of intracellular condensates.

许多细胞过程使用相分离将生物分子隔离到无膜细胞器中 或冷凝物。这个概念在细胞核中特别重要。大多数核凝聚物 与特定的染色质位点有关。因此，染色质组织和生物分子凝聚物 形成密切相关。拓扑学、表观遗传修饰或酶活性是可能的联系 染色质支架和蛋白质凝聚之间的联系。我们以前曾描述过冷凝物 将转录机制集中在超级增强子上， 和位点特异性特征，核浓缩物缺失。初步数据显示， 染色质装饰转录浓缩物的表面，低于常规的 显微镜利用最近的发展，在多路复用超分辨率显微镜， 染色质元件和蛋白质因子，我们将表征染色质富集的性质 层，并确定它是否反映了冷凝水表面作为一个积极的界面，调节 转录。我们将首次直接观察局部染色质拓扑结构的相互作用， 小鼠胚胎干中特定模型位点的表观遗传标记和冷凝物形成 细胞通过用基因组分辨率仔细检查基因座中的每个调控元件如何相互作用， 利用凝聚物，我们将确定染色质结构如增强子枢纽是否成核 缩合物，或者如果缩合物通过连接活性元件诱导特定的染色质拓扑结构。 最后，我们将在分子水平上研究冷凝物组分如何与 微环境和相关的染色质元素，并测试的假设，保留 转录因子增强结合位点占有率并稳定调节染色质处的缩合物。 有针对性的扰动和特定地点的观察将使我们能够回答这个问题，并开始 确定全基因组规则，预测精确染色质支持的缩合物形成。 这项研究计划的总体愿景是发展一个染色质组织的综合观点 和冷凝物形成。实验数据将激发新的概念框架， 将缩合物的全部复杂性整合到生物功能的范例中， 调控我们期待着新的生物物理学模型的自我组织和自我调节将出现 从我们对细胞内凝聚物的研究中