Roßmanith Johanna, Narberhaus Franz
Microbial Biology, Ruhr University Bochum, 44780 Bochum, Germany.
Microbial Biology, Ruhr University Bochum, 44780 Bochum, Germany
Nucleic Acids Res. 2016 Jun 20;44(11):5410-23. doi: 10.1093/nar/gkw232. Epub 2016 Apr 8.
Natural regulatory RNAs like riboswitches and RNA thermometers (RNAT) have considerable potential in synthetic biology. They are located in the 5' untranslated region (UTR) of bacterial mRNAs and sense small molecules or changes in temperature, respectively. While riboswitches act on the level of transcription, translation or mRNA stability, all currently known RNATs regulate translation initiation. In this study, we explored the modularity of riboswitches and RNATs and obtained regulatory devices with novel functionalities. In a first approach, we established three riboswitch-RNAT systems conferring dual regulation of transcription and translation depending on the two triggers ligand binding and temperature sensing. These consecutive fusions control gene expression in vivo and can even orchestrate complex cellular behavior. In another approach, we designed two temperature-controlled riboswitches by the integration of an RNAT into a riboswitch aptamer domain. These 'thermoswitches' respond to the cognate ligand at low temperatures and are turned into a continuous on-state by a temperature upshift. They represent the first RNATs taking control of transcription. Overall, this study demonstrates that riboswitches and RNATs are ideal for engineering synthetic RNA regulators due to their modular behavior.
像核糖开关和RNA温度计（RNAT）这样的天然调节性RNA在合成生物学中具有巨大潜力。它们分别位于细菌mRNA的5'非翻译区（UTR），可感应小分子或温度变化。虽然核糖开关作用于转录、翻译或mRNA稳定性水平，但目前已知的所有RNAT都调节翻译起始。在本研究中，我们探索了核糖开关和RNAT的模块化，并获得了具有新功能的调节装置。在第一种方法中，我们建立了三个核糖开关-RNAT系统，根据配体结合和温度感应这两种触发因素对转录和翻译进行双重调节。这些连续的融合体在体内控制基因表达，甚至可以协调复杂的细胞行为。在另一种方法中，我们通过将RNAT整合到核糖开关适体结构域中设计了两个温度控制的核糖开关。这些“热开关”在低温下对同源配体作出反应，并通过温度升高转变为持续的开启状态。它们代表了第一批控制转录的RNAT。总体而言，这项研究表明，由于其模块化行为，核糖开关和RNAT是工程化合成RNA调节器的理想选择。
