Engineering Multiplexed Inflammation Sensing Circuit for Gut Modulation

The gut contains trillions of microbes that interact with host cells and monitor changes in the environment. Opportunistic pathogens exploit environmental conditions to stimulate their growth and virulence, leading to relapsing chronic disorders such as inflammatory bowel disease (IBD) in the gut. Despite the severity of these disorders, current therapies are effective in less than 30% of patients due to the lack of adherence to prescription schedules and broad, off-target effects. Fortunately, synthetic biology offers exciting opportunities to engineer smart microbial therapeutics that may colonize the gut, providing in situ surveillance and disease modulation. However, many current engineered microbes have been engineered to respond only to single gut environmental factors. In probiotic E. coli strain Nissle 1917, we implement the previously characterized split activator AND logic gate. Our system is capable of multiplexing two input signals, selective activation to an inflammatory biomarker, tetrathionate, and secondary input signal IPTG. We report 4-6 fold induction with minimal leak when both activators are present. We model the dynamics of the AND gate using chemical reaction networks, and by tuning parameters in silico, we identified perturbations that affect selectivity of our circuit. We anticipate that our results will prove useful for designing living therapeutics for spatial targeting and signal processing in complex environments.