(157d) Engineering Multicellular Motility-Based Biosensors Systems

The ability to robustly control cellular motility in response to an external stimulus can facilitate the development of biosensors or the construction of micron-sized robots that use microbes to enable movement. By separating the sensing and actuating cells, and coordinating their behavior via cell-cell communication, we can improve system design through modularity, signal amplification and logic gate-type responses to multiple inputs. Our first goal was to use a synthetic biology approach to build a system where the motility of Escherichia coli is controlled by a cell-cell communication signal. Towards this goal, we have designed and constructed a system where E. coli motility is controlled by an acyl homoserine lactone (AHL) quorum sensing (QS) signal molecule.  Unlike previous efforts that used transcriptional activation-based QS systems to control motility, we used the repressor-based esa QS system to enable tight regulation of a key motility gene.  We have demonstrated that we can restore motility in an E. coli strain with a motA deletion by expressing motA under the control of the esa promoter, which is tightly repressed by the QS transcriptional regulator (EsaR) in the absence of AHL. Expression of motA is triggered by AHL, which caused EsaR to dissociate from the esa promoter. To quantify motility, we measured the swimming of cells through semi-solid agar plates. We observed that cells do not swim in the absence of AHL, and swimming is induced in the presence of micromolar concentrations of AHL. Overall, our engineered communication-dependent motility (CoMot) cells show AHL dependent motility. We replaced wild type EsaR with a mutant that responds to nanomolar AHL concentrations to increase the AHL sensitivity of the cells (CoMot+).  Interestingly, directional swimming of the CoMot and CoMot+ cells was observed up an AHL gradient even though chemotaxis was not directly controlled.  Further, we have demonstrated that the cells respond to AHL produced by a second strain of E. coli. When the AHL production is inducible, the combination of AHL-producers and the CoMot cells enable a novel biosensing framework. The CoMot strains engineered here can be used to build synthetic networks and biosensor systems where a motility output is tightly regulated and controlled by cell-cell communication.