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

Abstract

O_LISynthetic 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. C_LIO_LIWe have adapted bacterial allosteric transcription factors (aTFs) to control gene expression in plants in a ligand-specific manner. The aTFs used here function as transcription repressors of semi-synthetic promoters, and aTF activity is regulated by specific plant metabolites, especially phenylpropanoid-related molecules. Using these aTFs, we also designed synthetic genetic circuits capable of computing Boolean logic operations. C_LIO_LIThree aTFs, CouR, FapR and TtgR, were able to achieve [~]95% repression of their respective target promoters. For TtgR, a 6-fold de-repression could be triggered by inducing its ligand (naringenin) 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. C_LIO_LIWe showed that biosensors can be implemented in plants to detect phenylpropanoid-related metabolites and activate a genetic circuit that follows a pre-defined logic, demonstrating their potential as tools for exerting control over plant metabolic pathways and facilitating the bioproduction of natural products. C_LI