Genetically encoded Boolean logic operators to sense and integrate phenylpropanoid metabolite levels in plants.

ABSTRACT

Synthetic biology has the potential to revolutionize biotechnology, public health, and agriculture. Recent studies have shown the enormous potential of plants as chassis for synthetic biology applications. However, tools to precisely manipulate metabolic pathways for bioproduction in plants are still needed. We used bacterial allosteric transcription factors (aTFs) that control gene expression in a ligand-specific manner and tested their ability to repress semi-synthetic promoters in plants. We also tested the modulation of their repression activity in response to specific plant metabolites, especially phenylpropanoid-related molecules. Using these aTFs, we also designed synthetic genetic circuits capable of computing Boolean logic operations. Three aTFs, CouR, FapR, and TtgR, achieved c. 95% repression of their respective target promoters. For TtgR, a sixfold de-repression could be triggered by inducing its ligand accumulation, showing its use as biosensor. Moreover, we designed synthetic genetic circuits that use AND, NAND, IMPLY, and NIMPLY Boolean logic operations and integrate metabolite levels as input to the circuit. We showed that biosensors can be implemented in plants to detect phenylpropanoid-related metabolites and activate a genetic circuit that follows a predefined logic, demonstrating their potential as tools for exerting control over plant metabolic pathways and facilitating the bioproduction of natural products.