Tracking regulatory DNA in action Cis-regulatory DNA elements such as enhancers and promoters are critical for transcription regulation. Little is known about the relationship between these elements' transcriptional activity and their mobility within the nucleus in living cells. Gu et al. developed a strategy to deliver multiple RNAs to guide inactive Cas9 to label these elements. Quantitative measurement of their movement during stem cell differentiation revealed that increased DNA loci mobility correlated with transcriptional activation. Science, this issue p. 1050 Live-cell imaging of cis-regulatory DNA elements reveals an intrinsic connection between their transcriptional activity and nuclear mobility. To achieve guide RNA (gRNA) multiplexing and an efficient delivery of tens of distinct gRNAs into single cells, we developed a molecular assembly strategy termed chimeric array of gRNA oligonucleotides (CARGO). We coupled CARGO with dCas9 (catalytically dead Cas9) imaging to quantitatively measure the movement of enhancers and promoters that undergo differentiation-associated activity changes in live embryonic stem cells. Whereas all examined functional elements exhibited subdiffusive behavior, their relative mobility increased concurrently with transcriptional activation. Furthermore, acute perturbation of RNA polymerase II activity can reverse these activity-linked increases in loci mobility. Through quantitative CARGO-dCas9 imaging, we provide direct measurements of cis-regulatory element dynamics in living cells and distinct cellular and activity states and uncover an intrinsic connection between cis-regulatory element mobility and transcription.

中文翻译：

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哺乳动物顺式调控元件核迁移率的转录耦合变化


追踪调控 DNA 的作用 顺式调控 DNA 元件（例如增强子和启动子）对于转录调控至关重要。人们对这些元件的转录活性与其在活细胞核内的迁移性之间的关系知之甚少。顾等人。开发了一种策略来传递多个 RNA 来引导非活性 Cas9 来标记这些元素。对干细胞分化过程中它们运动的定量测量表明，DNA 位点移动性的增加与转录激活相关。科学，本期第 14 页。 1050 顺式调控 DNA 元件的活细胞成像揭示了它们的转录活性和核迁移率之间的内在联系。为了实现引导 RNA (gRNA) 多重化以及将数十种不同的 gRNA 有效递送到单细胞中，我们开发了一种称为 gRNA 寡核苷酸嵌合阵列 (CARGO) 的分子组装策略。我们将 CARGO 与 dCas9（催化死亡 Cas9）成像结合起来，定量测量活胚胎干细胞中经历分化相关活性变化的增强子和启动子的运动。尽管所有检查的功能元件都表现出亚扩散行为，但它们的相对移动性随着转录激活而同时增加。此外，RNA 聚合酶 II 活性的急性扰动可以逆转这些与活性相关的基因座迁移率的增加。通过定量 CARGO-dCas9 成像，我们可以直接测量活细胞中的顺式调控元件动态以及不同的细胞和活动状态，并揭示顺式调控元件迁移性和转录之间的内在联系。