Construction and Characterization of Consensus Toggle Switches

A genetic toggle switch is a circuit that possesses two stable opposite states of gene expression. Many synthetic circuits that are applied to biotechnology are based on toggle switches, such as biosensors. However, extrinsic and intrinsic noise can cause random switching and generate a variety of gene expression states. To make circuits more robust, we propose the creation of a consensus toggle switch that will have the ability to amplify induction signals among neighboring cells and achieve a consensus in the population. We will create the consensus by employing quorum sensing in our circuits. Quorum sensing (QS) is the mechanism through which bacteria communicate with each other, by producing signaling molecules that can affect gene expression in a density-dependent way. We hypothesize that synthetic toggles that coordinate their behavior with QS can better perform tasks that are difficult for uncoordinated systems. Here, we constructed 4 versions of consensus toggles to understand how the presence/absence of positive feedback loops in quorum-sensing genes and direct/indirect repression affect toggle behavior. In our designs, each possible state of the toggle is reported by one color: yellow (YFP) or blue (CFP). We find that the dynamics of consensus toggle switches depend on their regulatory topologies. Topologies that contain indirect repression show a peak of YFP expression followed by a shutdown, while topologies with direct repression do not shut down. Furthermore, topologies with indirect repression do not show distinguishable CFP expression between induced and uninduced samples, while topologies with direct repression do. Topologies that lack extra feedback loops show less variability, while topologies that contain these extra feedback loops are more variable. These results enable the selection of a topology that works better for the creation of a consensus toggle. In the microscope, the toggle with direct repression and without extra feedback loops showed faster switch compared to a conventional toggle. These results are encouraging, because a consensus toggle has the potential of generating more sensitive and robust detection of signals than regular toggles. And to further apply this circuit, such as for the detection of diseases/infections in the mammalian gastrointestinal tract or for the surveillance and bioremediation of environments, it is desired that the circuit shows sensitivity and robustness.