(796b) Construction of a Cell-Cell Communication-Dependent and Gate

The integration of cell-cell communication signals with other chemical inputs at a single promoter has the potential to provide a new set of tools for applications that require cells to respond to both population density and the presence of a second, exogenous signal. We have contstructed and characterized a set of promoters that behave as AND-gates, where gene expression is turned on only in the presence of a quorum-sensing (QS) signal and an exogenous inducer. In addition to widely used transcription factor operator pairs (LacI/LacO, TetR/TetO), we have employed the EsaR protein and its DNA target, the esa box. Unlike LuxR and other common QS regulators, EsaR can act as a repressor, where binding an acyl-homoserine lactone (AHL) molecule causes it to release its DNA target. We have shown that expression from the AND-gate promoters occurs only in the presence of both AHL and either anhydrotetracycine or IPTG. Enabling endogenous AHL production via the expression of an AHL synthase, EsaI, allowed us to control gene expression in a density-dependent manner, but only in the presence of the exogenous inducer. This system provides a unique alternative to typical autoinduction systems and provides a framework for the generation of AND-gates that respond to both cell-cell communication and any metabolite for which there is an available regulator.