The ultimate goal of synthetic biology is to construct artificial cells, aka minimal cells. Such artificial cells have high clinical relevance, with potential applications ranging from gene therapy, synthetic oxygen carrier, to drug delivery. By far, the creation of a completely artificial and fully functional cell has not yet been successful. One of the main hurdles of artificial cell construction is to achieve a comprehensive regulation of information. RNA can directly produce protein and possess broad interactions with non-genetic components; thus, RNA has been considered suitable information carrier for artificial cells as their DNA counterpart. However, current synthetic RNA tools with regulatory functions, such as mRNA switches and mRNA circuits, all act in a simple binary manne...[ Read more ]

The ultimate goal of synthetic biology is to construct artificial cells, aka minimal cells. Such artificial cells have high clinical relevance, with potential applications ranging from gene therapy, synthetic oxygen carrier, to drug delivery. By far, the creation of a completely artificial and fully functional cell has not yet been successful. One of the main hurdles of artificial cell construction is to achieve a comprehensive regulation of information. RNA can directly produce protein and possess broad interactions with non-genetic components; thus, RNA has been considered suitable information carrier for artificial cells as their DNA counterpart. However, current synthetic RNA tools with regulatory functions, such as mRNA switches and mRNA circuits, all act in a simple binary manner (having only ON or OFF stages of protein signal readout), which are insufficient for the need of building regulatory networks. In this study, we innovatively engineered two novel types of advanced mRNA circuits. An engineered regulatory domain enables a single mRNA circuit to sense two different types of input molecules in a competitive manner. A smart design allows the same input molecules to generate opposite effects on two mRNA switches, affording a dual-output mRNA circuit. These two circuits can both be easily adapted to sense other input molecules of the same category, and can be flexibly integrated with other mRNA circuits to build complex RNA circuits. We believe the two novel mRNA circuits have strong potential for biomedical application. Moreover, the smart design demonstrated in this work also sheds light on the engineering of other mRNA circuits, and eventually the building of RNA-based artificial cells.