The 3D-Structures of the Pre-Pro-B and Pro-B Cell Genomes

描述(由申请人提供)：最近的研究提供了大量证据表明，基因组不仅由线性结构组成，而且还被折叠成精细的模式，允许被大基因组距离分开的基因组元素之间的相互作用。基因组的精确折叠和拓扑谱仍有待确定。最近的研究为免疫球蛋白重链(IgH)和球蛋白基因座的三维结构提供了洞察力。这些数据表明，这些基因座被组织成多个以环束为特征的亚室，这与多环-亚室(MLS)模型一致。我们现在面临的问题是，整个基因组是否像MLS模型所提出的那样折叠。为了解决这个问题，我们最近采用了甲醛交联法，允许鉴定前B细胞中相互作用的基因组元件的全基因组网络。我们的初步数据表明，正如MLS模型预测的那样，前B细胞基因组确实被组织成环簇，这些环簇通过连接子连接在一起。在这里，我们建议继续进行这些研究。我们将对个体隔间进行深入的分析，并检查它们是如何组织的，以及它们与B系特定的基因表达程序是如何关联的。我们将识别和描述染色体内的相互作用。绝缘体和桥联因素将从生化和功能的角度进行识别和表征。最后，我们建议确定前-前-B和前-B细胞中有丝分裂染色体的结构。总体目标将是破译前-B和前-B细胞染色质纤维形成的构象和轨迹的光谱，以及染色质结构如何与功能相关。 公共卫生相关性：这里描述的建议旨在开发一种新的方法来描述Pre-proB和Pro-B细胞中染色质纤维的折叠。这项研究将提供对允许增强子和启动子元件以适当的频率相互作用以调节下游靶基因表达的机制的物理见解。这些研究还可能提供对支撑基因组不稳定的机制的洞察。

DESCRIPTION (provided by applicant): Recent studies have provided substantial evidence that the genome consists not merely of linear structures but rather it is folded into elaborate patterns that permits interactions between genomic elements separated by large genomic distances. The precise folding and the spectrum of topologies of the genome remain to be determined. Recent studies have provided insight into the 3D-architectures of the immunoglobulin heavy chain (Igh) and ¿-globin loci. These data have revealed that these loci are organized into multiple compartments that are characterized by bundles of loops, consistent with the Multiple-Loop-Subcompartment (MLS) Model. We are now faced with the question as to whether the entire genome is folded as proposed by the MLS model. To address this question, we have recently employed a formaldehyde cross-linking approach that permits the identification of a genome-wide network of interacting genomic elements in pro-B cells. Our preliminary data indicates that the pro-B cell genome indeed is organized into clusters of loops that are connected by linkers as predicted by the MLS model. Here we would propose to continue these studies. We would perform an in depth analysis of individual compartments and examine how they are organized and how they relate to B-lineage specific programs of gene expression. We would identify and characterize intra-chromosomal interactions. Insulators and bridging factors would be identified and characterized both from a biochemical and functional perspective. Finally, we propose to determine the structure of mitotic chromosomes in pre-pro-B and pro-B cells. The overall goal would be to decipher the spectrum of conformations and trajectories formed by the pre-pro-B and pro-B cell chromatin fibers and how chromatin structure relates to function. PUBLIC HEALTH RELEVANCE: The proposal described here is aimed at developing a novel approach to describe the folding of the chromatin fiber in pre-pro-B and pro-B cells. The study would provide physical insight into the mechanism that permits enhancer and promoter elements to interact with each other with the appropriate frequencies in order to modulate down- stream target gene expression. The studies may also provide insight into the mechanisms that underpin genomic instability.