(56a) Synthetic Autoinducer-2 Triggered Expression for Quorum Sensing Surveillance

Bacterial multicellularity is primarily coordinated by a communicative process known as quorum sensing (QS).  Its signal-based and “switch-like” regulation of phenotypic changes provides an inspiring framework for interaction-driven expression among distinct populations within a complex bacterial community.  Accordingly, we seek to emulate the QS “switch” for useful purposes including controlled protein expression.¹ In the current work, we have built on this theme to use bacteria to report on their nearest neighbors and on their immediate environment. We demonstrate expression of streptavidin-binding peptide as a response to the presence of autoinducer-2, a QS signaling molecule.  The engineered cell surveys its environment for AI-2, and upon recognition of the signal, the cell then expresses a surface-displayed streptavidin binding peptide (SBP), rendering it readily susceptible for concentration, separation, and amplified detection. This strategy expands upon previous work that demonstrated the ability of an operon under AI-2 regulation to be rewired within a dual plasmid system for the fluorescent reporting of AI-2.  Here, we have associated SBP expression with fluorescence- both by labeling the cell surface with an affinity tag and by intrinsically expressing a fluorescent protein.  In this way, the bacteria synthetically feature a surveillance mechanism to interact amongst a quorum sensing population yet remain fluorescently distinguishable, to sense and respond to the event of AI-2 exchange, and to be recovered.  Our data characterizes each of these features.  Furthermore, we have engineered the sensitivity of the surveyor cells by tuning the QS “switch” at specific thresholds of autoinducer signaling.   Our surveillance mechanism uniquely capitalizes on the event of QS to achieve selective separation from a mixed, larger community. In this way, cell-based surveillance could be applicable as an alternative to current detection methods for bacterial contamination, especially that which is pathogenic. Benefit is provided by the sensitive molecular recognition capability per single surveyor cell.  Additionally, concentration of fluorescent output via cellular affinity to streptavidin enables an amplification of the original signal and further enhances detection ability.   Overall, this work is representative of a fertile opportunity for engineers to interrogate and manipulate cell behavior by eavesdropping on cell-to-cell communication at the level of individual cells.

¹Tsao et al., Metab. Eng., 12, 291 (2010)