Department of Physics, Massachusetts Institute of Technology, Cambridge, MA, USA.
Department of Media Arts and Sciences, Massachusetts Institute of Technology, Cambridge, MA, USA.
Nat Biotechnol. 2021 Mar;39(3):320-325. doi: 10.1038/s41587-020-0704-z. Epub 2020 Oct 19.
Current approaches to single-cell RNA sequencing (RNA-seq) provide only limited information about the dynamics of gene expression. Here we present RNA timestamps, a method for inferring the age of individual RNAs in RNA-seq data by exploiting RNA editing. To introduce timestamps, we tag RNA with a reporter motif consisting of multiple MS2 binding sites that recruit the adenosine deaminase ADAR2 fused to an MS2 capsid protein. ADAR2 binding to tagged RNA causes A-to-I edits to accumulate over time, allowing the age of the RNA to be inferred with hour-scale accuracy. By combining observations of multiple timestamped RNAs driven by the same promoter, we can determine when the promoter was active. We demonstrate that the system can infer the presence and timing of multiple past transcriptional events. Finally, we apply the method to cluster single cells according to the timing of past transcriptional activity. RNA timestamps will allow the incorporation of temporal information into RNA-seq workflows.
目前的单细胞 RNA 测序 (RNA-seq) 方法仅提供关于基因表达动态的有限信息。在这里，我们提出了 RNA 时间戳，这是一种通过 RNA 编辑推断 RNA-seq 数据中单个 RNA 年龄的方法。为了引入时间戳，我们使用由多个 MS2 结合位点组成的报告基因构建体标记 RNA，该构建体招募与 MS2 衣壳蛋白融合的腺苷脱氨酶 ADAR2。ADAR2 与标记的 RNA 结合会导致 A 到 I 的编辑随时间积累，从而可以以小时级的精度推断 RNA 的年龄。通过结合由同一启动子驱动的多个标记 RNA 的观察结果，我们可以确定启动子何时处于活跃状态。我们证明该系统可以推断过去多个转录事件的存在和时间。最后，我们将该方法应用于根据过去转录活性的时间对单细胞进行聚类。RNA 时间戳将允许将时间信息纳入 RNA-seq 工作流程。
