Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115, USA.
Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115, USA; Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
Cell. 2014 Nov 6;159(4):925-39. doi: 10.1016/j.cell.2014.10.002. Epub 2014 Oct 23.
Efforts to construct synthetic networks in living cells have been hindered by the limited number of regulatory components that provide wide dynamic range and low crosstalk. Here, we report a class of de-novo-designed prokaryotic riboregulators called toehold switches that activate gene expression in response to cognate RNAs with arbitrary sequences. Toehold switches provide a high level of orthogonality and can be forward engineered to provide average dynamic range above 400. We show that switches can be integrated into the genome to regulate endogenous genes and use them as sensors that respond to endogenous RNAs. We exploit the orthogonality of toehold switches to regulate 12 genes independently and to construct a genetic circuit that evaluates 4-input AND logic. Toehold switches, with their wide dynamic range, orthogonality, and programmability, represent a versatile and powerful platform for regulation of translation, offering diverse applications in molecular biology, synthetic biology, and biotechnology.
在活细胞中构建人工合成网络的努力受到了限制，因为能够提供宽动态范围和低串扰的调控组件的数量有限。在这里，我们报告了一类新设计的原核核糖调控因子，称为臂环开关，它可以响应具有任意序列的同源 RNA 来激活基因表达。臂环开关提供了高度的正交性，可以进行正向工程设计，以提供超过 400 的平均动态范围。我们表明，开关可以整合到基因组中，以调节内源性基因，并将其用作响应内源性 RNA 的传感器。我们利用臂环开关的正交性来独立调节 12 个基因，并构建了一个评估 4 输入与非门逻辑的遗传电路。臂环开关具有宽的动态范围、正交性和可编程性，代表了一种用于翻译调控的多功能和强大平台，在分子生物学、合成生物学和生物技术中具有广泛的应用。
