The manipulation of quorum sensing (QS) molecules to guide bacterial behavior is a promising approach to applications from protein engineering to human disease. However, there has been little progress in developing controller cells that modulate phenotype-directing metabolic cues of the coordinated system. In this work, we characterized the kinetics of the lsr operon in E. coli that produces and responds to the 'universal' quorum sensing signal, autoinducer-2 (AI-2). This was accomplished through the generation of a suite of bacterial strains that overexpress the respective components of AI-2 processing: uptake (LsrACDB), phosphorylation (LsrK) and degradation (LsrFG). These engineered ‘bacterial vacuums’ show the ability to regulate the extracellular AI-2 environment, silence bacterial communication ( &gt; 90 % reduction in coordinated gene expression), and disturb biofilm formation (~20% reduction in biofilm mass) without exerting selective pressure. Building on this system, we then inserted a QS-responsive switch, thereby decoupling the need for exogenous addition of an inducing agent (i.e. IPTG), which then activates the ‘bacterial vacuum’ only at sufficient QS thresholds. Using the insight gained from our mathematical model of these AI-2 processing mechanisms (Fig 1A), we enhanced the sensitivity of this QS switch through AI-2 activated expression of the uptake (LsrACDB) mechanism. Finally, we show that the synthetic development of bacteria with varied sensitivities to QS levels can not only interrogate environments to detect bacteria, but can also produce self-patterned gene expression in co-cultures dependent only on QS levels(Fig 1B). We suggest that similarly engineered commensal E. coli may provide a means for altering behavior in the gut microbiome. Alternatively, by manipulating the signal transduction process, we can swap the output production of co-culture systems from one product to another.

Autonomous ‘bacterial vacuums’ that control communication and enable tunable gene expression

249th National Meeting (2015)

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