Regulation of chromatin folding in space and time

原始项目摘要 真核基因组以其线性序列编码遗传信息，但适当地表达 他们的基因需要染色体折叠成复杂的、空间上不同的三维结构。 基因组方法的最新进展揭示了DNA相互作用的层次结构，从小到小 将基因和增强子连接到更大的染色体区域和核的染色质环 车厢。然而，尽管这些组织特征和它们的 对基因功能的影响，我们对染色体是如何在空间上分割的了解非常有限， 功能包装，并相对定位在细胞核内。技术限制也阻碍了我们的 能够提出关于细胞间的可变性和染色质折叠之间的关系的问题， 定位，并在单个单元分辨率下工作。 我们之前的研究包括开发两种使用原位荧光的技术 杂交(FISH)在单细胞分辨率下询问染色体定位。我们的目标是在此基础上再接再厉 研究并使用这些工具来阐明染色体片段是如何找到彼此并形成稳定的 细胞内的相互作用。我可以设想我们的工作立即进入三个阶段。首先是开发一种快速和 精确识别参与染色体相互作用的候选者的方法。第二个是建立一个 一组基于现场的分析，可以用来表征候选人的特征，第三个是翻译我们的 从模型生物到人类的发现。总的来说，这里提出的研究将揭示新的分子 核组织的潜在机制，为研究染色质折叠和 定位是建立和遗传的，以及组织功能失调如何导致疾病。

Original Project Abstract Eukaryotic genomes encode genetic information in their linear sequence, but appropriate expression of their genes requires chromosomes to fold into complex and spatially distinct three-dimensional structures. Recent advances in genomic-based approaches have uncovered a hierarchy of DNA interactions, from small chromatin loops that connect genes and enhancers to larger chromosomal domains and nuclear compartments. However, despite the remarkable conservation of these organizational features and their impact on gene function, we have a very limited understanding of how chromosomes are spatially partitioned, functionally packaged, and relatively positioned in the nucleus. Technical limitations have also hindered our ability to ask questions regarding cell-to-cell variability and the relationship between chromatin folding, positioning, and function at single cell resolution. Our previous studies involved the development of two technologies that use fluorescent in situ hybridization (FISH) to interrogate chromosome positioning at single-cell resolution. Our goal is to build on this work and use these tools to elucidate how chromosomal segments find each other and then form stable interactions within cells. I can envision three immediate stages for our work. The first is developing a rapid and precise method for identifying candidates involved in chromosome interactions. The second is establishing a battery of in situ-based assays that can be used to characterize the candidates, and the third is translating our findings from model organisms to humans. Collectively, the studies proposed here will uncover novel molecular mechanisms underlying nuclear organization, providing a new avenue to study how chromatin folding and positioning is established and inherited, and how dysfunctional organization contributes to disease.