Individual elements within a superenhancer can act in a cooperative or temporal manner, but the underlying mechanisms remain obscure. We recently identified an Irf8 superenhancer, within which different elements act at distinct stages of type 1 classical dendritic cell (cDC1) development. The +41-kb Irf8 enhancer is required for pre-cDC1 specification, while the +32-kb Irf8 enhancer acts to support subsequent cDC1 maturation. Here, we found that compound heterozygous Δ32/Δ41 mice, lacking the +32- and +41-kb enhancers on different chromosomes, show normal pre-cDC1 specification but, surprisingly, completely lack mature cDC1 development, suggesting cis dependence of the +32-kb enhancer on the +41-kb enhancer. Transcription of the +32-kb Irf8 enhancer-associated long noncoding RNA (lncRNA) Gm39266 is also dependent on the +41-kb enhancer. However, cDC1 development in mice remained intact when Gm39266 transcripts were eliminated by CRISPR/Cas9-mediated deletion of lncRNA promoters and when transcription across the +32-kb enhancer was blocked by premature polyadenylation. We showed that chromatin accessibility and BATF3 binding at the +32-kb enhancer were dependent on a functional +41-kb enhancer located in cis. Thus, the +41-kb Irf8 enhancer controls the subsequent activation of the +32-kb Irf8 enhancer in a manner that is independent of associated lncRNA transcription.

中文翻译：

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Irf8 位点中的顺式相互作用调节阶段依赖性增强子激活


超级增强器中的各个元件可以以合作或暂时的方式起作用，但潜在的机制仍然模糊。我们最近发现了一种Irf8超级增强子，其中不同的元件在 1 型经典树突状细胞 (cDC1) 发育的不同阶段发挥作用。 +41-kb Irf8增强器是 cDC1 之前规范所必需的，而 +32-kb Irf8增强器则用于支持后续的 cDC1 成熟。在这里，我们发现复合杂合的Δ32/Δ41小鼠在不同染色体上缺乏+32-和+41-kb增强子，表现出正常的cDC1前规范，但令人惊讶的是，完全缺乏成熟的cDC1发育，表明+32的顺式依赖性+41-kb 增强子上的 -kb 增强子。 +32-kb Irf8增强子相关的长非编码 RNA (lncRNA) Gm39266 的转录也依赖于 +41-kb 增强子。然而，当 CRISPR/Cas9 介导的 lncRNA 启动子删除消除了 Gm39266 转录本，并且当 +32-kb 增强子的转录被过早的聚腺苷酸化阻断时，小鼠中的 cDC1 发育保持完整。我们表明，+32-kb 增强子处的染色质可及性和 BATF3 结合取决于位于cis的功能性 +41-kb 增强子。因此，+41-kb Irf8增强子以独立于相关lncRNA转录的方式控制+32-kb Irf8增强子的后续激活。