The Mediator complex regulates enhancer-promoter interactions [Tiled-Micro-Capture-C]. The Mediator complex regulates enhancer-promoter interactions [Tiled-Micro-Capture-C]

Enhancer-mediated gene activation generally requires physical proximity between enhancers and their target gene promoters. However, the molecular mechanisms by which interactions between enhancers and promoters are formed are not well understood. Here, we investigate the function of the Mediator complex in the regulation of enhancer-promoter interactions, by combining rapid protein depletion and high-resolution MNase-based chromosome conformation capture approaches. We show that depletion of Mediator leads to reduced enhancer-promoter interaction frequencies, which are associated with a strong decrease in gene expression. In addition, we find increased interactions between CTCF-binding sites upon Mediator depletion. These changes in chromatin architecture are associated with a re-distribution of the Cohesin complex on chromatin and a reduction in Cohesin occupancy specifically at enhancers. Our results indicate that enhancer-promoter interactions are dependent on an interplay between the Mediator and Cohesin complexes and provide new insights into the molecular mechanisms by which communication between enhancers and promoters is regulated. Overall design: Tiled-Micro-Capture-C (Tiled-MCC). Details are described in the Methods section of the paper.

增强子介导的基因激活通常需要增强子与其靶基因启动子之间形成物理邻近关系。然而，增强子与启动子之间相互作用的形成机制目前尚不清楚。本研究结合快速蛋白耗竭技术与高分辨率微球菌核酸酶（MNase）依赖型染色体构象捕获方法，探究中介体复合物（Mediator complex）在调控增强子-启动子相互作用中的功能。研究结果显示，中介体复合物的耗竭会降低增强子-启动子的相互作用频率，这与基因表达的显著下调密切相关。此外，我们发现中介体复合物耗竭后，CTCF结合位点（CTCF-binding sites）之间的相互作用有所增强。染色质结构的这些变化与黏连蛋白复合物（Cohesin complex）在染色质上的重新分布，以及其在增强子区域的结合占有率特异性降低密切相关。本研究结果表明，增强子-启动子相互作用依赖于中介体复合物与黏连蛋白复合物之间的相互调控，并为增强子与启动子之间通信调控的分子机制提供了新的见解。整体实验设计：平铺式微捕获-染色体构象捕获（Tiled-Micro-Capture-C, Tiled-MCC），具体细节详见论文的方法部分。