Remote enhancers are thought to interact with their target promoters via physical proximity, yet the importance of this proximity for enhancer function remains unclear. Here we investigate the three-dimensional (3D) conformation of enhancers during mammalian development by generating high-resolution tissue-resolved contact maps for nearly a thousand enhancers with characterized in vivo activities in ten murine embryonic tissues. Sixty-one percent of developmental enhancers bypass their neighboring genes, which are often marked by promoter CpG methylation. The majority of enhancers display tissue-specific 3D conformations, and both enhancer–promoter and enhancer–enhancer interactions are moderately but consistently increased upon enhancer activation in vivo. Less than 14% of enhancer–promoter interactions form stably across tissues; however, these invariant interactions form in the absence of the enhancer and are likely mediated by adjacent CTCF binding. Our results highlight the general importance of enhancer–promoter physical proximity for developmental gene activation in mammals.

远程增强子被认为通过物理接近性与其目标启动子相互作用，但这种接近性对于增强子功能的重要性仍不清楚。在这里，我们通过为近千个增强子生成高分辨率的组织分辨接触图来研究增强子在哺乳动物发育过程中的三维（3D）构象，这些增强子在十个小鼠胚胎组织中具有体内活性特征。61% 的发育增强子绕过其邻近基因，这些基因通常以启动子 CpG 甲基化为标志。大多数增强子显示出组织特异性的 3D 构象，并且增强子-启动子和增强子-增强子相互作用在增强子体内激活时适度但持续增加。不到 14% 的增强子-启动子相互作用在组织中稳定形成；然而，这些不变的相互作用是在没有增强子的情况下形成的，并且可能是由相邻的 CTCF 结合介导的。我们的结果强调了增强子-启动子物理接近性对于哺乳动物发育基因激活的普遍重要性。