School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, New York, USA.
Nat Chem Biol. 2015 Mar;11(3):214-20. doi: 10.1038/nchembio.1737. Epub 2015 Feb 2.
We expanded the mechanistic capability of small RNAs by creating an entirely synthetic mode of regulation: small transcription activating RNAs (STARs). Using two strategies, we engineered synthetic STAR regulators to disrupt the formation of an intrinsic transcription terminator placed upstream of a gene in Escherichia coli. This resulted in a group of four highly orthogonal STARs that had up to 94-fold activation. By systematically modifying sequence features of this group, we derived design principles for STAR function, which we then used to forward engineer a STAR that targets a terminator found in the Escherichia coli genome. Finally, we showed that STARs could be combined in tandem to create previously unattainable RNA-only transcriptional logic gates. STARs provide a new mechanism of regulation that will expand our ability to use small RNAs to construct synthetic gene networks that precisely control gene expression.
我们通过创建一种完全合成的调控模式——小转录激活 RNA（STARs），扩展了小 RNA 的机制能力。我们使用两种策略，设计了合成 STAR 调节剂来破坏放置在大肠杆菌中基因上游的固有转录终止子的形成。这导致了一组四个高度正交的 STAR，其激活倍数高达 94 倍。通过系统地修饰这个星团的序列特征，我们得出了 STAR 功能的设计原则，然后我们用这些原则来正向设计一个针对大肠杆菌基因组中发现的终止子的 STAR。最后，我们表明 STARs 可以串联组合，形成以前无法实现的仅 RNA 转录逻辑门。STARs 提供了一种新的调控机制，将扩大我们使用小 RNA 构建精确控制基因表达的合成基因网络的能力。
