Center for Genomic Regulation (CRG), Barcelona Institute of Science and Technology, Barcelona, Spain.
Institute for Research in Biomedicine (IRB Barcelona), Barcelona Institute of Science and Technology, Barcelona, Spain.
Nat Struct Mol Biol. 2022 Oct;29(10):1011-1023. doi: 10.1038/s41594-022-00839-y. Epub 2022 Oct 11.
The linear sequence of DNA provides invaluable information about genes and their regulatory elements along chromosomes. However, to fully understand gene function and regulation, we need to dissect how genes physically fold in the three-dimensional nuclear space. Here we describe immuno-OligoSTORM, an imaging strategy that reveals the distribution of nucleosomes within specific genes in super-resolution, through the simultaneous visualization of DNA and histones. We combine immuno-OligoSTORM with restraint-based and coarse-grained modeling approaches to integrate super-resolution imaging data with Hi-C contact frequencies and deconvoluted micrococcal nuclease-sequencing information. The resulting method, called Modeling immuno-OligoSTORM, allows quantitative modeling of genes with nucleosome resolution and provides information about chromatin accessibility for regulatory factors, such as RNA polymerase II. With Modeling immuno-OligoSTORM, we explore intercellular variability, transcriptional-dependent gene conformation, and folding of housekeeping and pluripotency-related genes in human pluripotent and differentiated cells, thereby obtaining the highest degree of data integration achieved so far to our knowledge.
线性 DNA 序列为基因及其在染色体上的调控元件提供了宝贵的信息。然而，要全面了解基因的功能和调控，我们需要剖析基因在三维核空间中的物理折叠方式。在这里，我们描述了 immuno-OligoSTORM，这是一种成像策略，通过同时可视化 DNA 和组蛋白，以超分辨率揭示特定基因内核小体的分布。我们将 immuno-OligoSTORM 与基于约束和粗粒化建模方法相结合，将超分辨率成像数据与 Hi-C 接触频率和去卷积微球菌核酸酶测序信息整合在一起。该方法称为 Modeling immuno-OligoSTORM，可对具有核小体分辨率的基因进行定量建模，并为 RNA 聚合酶 II 等调控因子提供有关染色质可及性的信息。利用 Modeling immuno-OligoSTORM，我们探索了人类多能性和分化细胞中细胞间变异性、转录依赖性基因构象以及管家基因和多能性相关基因的折叠，从而获得了迄今为止我们所知的最高程度的数据集成。
