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1. Antiactivators prevent self-sensing in Pseudomonas aeruginosa quorum sensing

   https://doi.org/10.1073/pnas.2201242119  

   Smith, Parker; Schuster, Martin (June 2022, Proceedings of the National Academy of Sciences)

   Quorum sensing is described as a widespread cell density-dependent signaling mechanism in bacteria. Groups of cells coordinate gene expression by secreting and responding to diffusible signal molecules. Theory, however, predicts that individual cells may short-circuit this mechanism by directly responding to the signals they produce irrespective of cell density. In this study, we characterize this self-sensing effect in the acyl-homoserine lactone quorum sensing system of Pseudomonas aeruginosa . We show that antiactivators, a set of proteins known to affect signal sensitivity, function to prevent self-sensing. Measuring quorum-sensing gene expression in individual cells at very low densities, we find that successive deletion of antiactivator genes qteE and qslA produces a bimodal response pattern, in which increasing proportions of constitutively induced cells coexist with uninduced cells. Comparing responses of signal-proficient and -deficient cells in cocultures, we find that signal-proficient cells show a much higher response in the antiactivator mutant background but not in the wild-type background. Our results experimentally demonstrate the antiactivator-dependent transition from group- to self-sensing in the quorum-sensing circuitry of P. aeruginosa . Taken together, these findings extend our understanding of the functional capacity of quorum sensing. They highlight the functional significance of antiactivators in the maintenance of group-level signaling and experimentally prove long-standing theoretical predictions. 

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2. Dynamic Control of Gene Expression with Riboregulated Switchable Feedback Promoters

   https://doi.org/10.1021/acssynbio.1c00015  

   Glasscock, Cameron J.; Biggs, Bradley W.; Lazar, John T.; Arnold, Jack H.; Burdette, Lisa A.; Valdes, Aliki; Kang, Min-Kyoung; Tullman-Ercek, Danielle; Tyo, Keith E.; Lucks, Julius B. (April 2021, ACS Synthetic Biology)

   null (Ed.)

   One major challenge in synthetic biology is the deleterious impacts of cellular stress caused by expression of heterologous pathways, sensors, and circuits. Feedback control and dynamic regulation are broadly proposed strategies to mitigate this cellular stress by optimizing gene expression levels temporally and in response to biological cues. While a variety of approaches for feedback implementation exist, they are often complex and cannot be easily manipulated. Here, we report a strategy that uses RNA transcriptional regulators to integrate additional layers of control over the output of natural and engineered feedback responsive circuits. Called riboregulated switchable feedback promoters (rSFPs), these gene expression cassettes can be modularly activated using multiple mechanisms, from manual induction to autonomous quorum sensing, allowing control over the timing, magnitude, and autonomy of expression. We develop rSFPs in Escherichia coli to regulate multiple feedback networks and apply them to control the output of two metabolic pathways. We envision that rSFPs will become a valuable tool for flexible and dynamic control of gene expression in metabolic engineering, biological therapeutic production, and many other applications. 

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3. Population-level bistability in Pseudomonas aeruginosa quorum sensing

   https://doi.org/10.1128/mbio.01713-25  

   Pettit_Estell, Bryce M; Schuster, Martin (October 2025, mBio)

   Fuqua, Clay (Ed.)

   ABSTRACT Quorum sensing (QS) is a widespread signaling mechanism in bacteria that coordinates collective behaviors according to population density. A foundational assumption in this field is that QS functions as a gene expression switch that synchronizes responses at the population level. While some studies indeed report homogeneous on/off transitions, others report heterogeneity at the cellular level, challenging the canonical view. To provide a formal context for these observations, we examined QS behavior at physiological steady state in the model bacteriumPseudomonas aeruginosa. We measured the expression of a centrallas-system component, the positively autoregulated signal synthase genelasI, at the population and single-cell level. In support of the canonical view and predictions from mathematical modeling, we show that thelassystem exhibits population-level bistability, with the entire unimodal population of cells switching synchronously between two stable states. We also show that bistable state switching exhibits hysteresis, indicative of memory within the system, with induced cells maintaining activation at considerably lower densities than previously uninduced cells. We confirm these behaviors in a subset of otherlas-QS controlled genes. Our study experimentally proves population-level bistability as a central, emergent property in a native QS system, with implications for physiology, pathogenesis, and synthetic biology.IMPORTANCEThis research investigates quorum sensing (QS), a common bacterial communication mechanism that controls processes like virulence, biofilm formation, and microbial warfare. Our study experimentally proves the long-standing notion that native QS can function as a true genetic switch that synchronizes all-or-none responses in a population of cells. We employ the well-understood LasI/LasR QS system of the opportunistic pathogenPseudomonas aeruginosaas a model. We show that the switch is bistable, with stable on and off states, and that it is hysteretic, with a type of memory that makes state-switching dependent on the initial condition. These properties impart robustness and stability to environmental changes akin to cellular developmental pathways; they have general implications for infection and its control, as well as genetic circuit design. 

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4. A family of linear plasmid phages that detect a quorum-sensing autoinducer exists in multiple bacterial species

   https://doi.org/10.1128/mbio.02320-25  

   Santoriello, Francis J; Bassler, Bonnie L (January 2026, mBio)

   Hatfull, Graham F (Ed.)

   ABSTRACT Temperate phages oscillate between lysogeny, a genomic maintenance state within a bacterial host, and lytic replication, in which the host is killed, and newly made phage particles are released. Successful transmission to new hosts requires that temperate phages appropriately time their transitions from lysogeny to lysis. It is well understood that temperate phages trigger lysis upon detection of host cell stress. Understanding of the breadth of cues that induce lysis expanded with the discovery of phages carrying quorum-sensing receptor genes that promote lytic induction exclusively at high host cell density. Bacteria engage in a cell-cell communication process called quorum sensing, which relies on the production, release, accumulation, and group-wide detection of extracellular signal molecules called autoinducers. Bacteria use quorum sensing to monitor changes in population density and synchronize collective behaviors. The temperate phage VP882 (φVP882) encodes VqmAφ—a homolog of its host’s quorum-sensing receptor/transcription factor VqmA. VqmAφ allows φVP882 to detect the accumulation of the host autoinducer called DPO. Presumably, launching the lytic induction program at high host cell density maximizes φVP882 transmission to new hosts. Here, by mining sequence databases for linear plasmid phages, we identify VP882-like phages in multiple DPO-producing bacterial species isolated at diverse times and geographic locations. We show that the VqmAφ homologs can indeed detect DPO and, in response, activate the lytic pathway. Our findings indicate that φVP882 is a member of a family of globally dispersed temperate phages that possess the potential to respond to host quorum sensing. IMPORTANCEThe discovery of quorum-sensing responsive linear plasmid phages has transformed understanding of phage-bacterial interactions by demonstrating inter-domain chemical communication. To date, however, examples of quorum-sensing responsive phages have been sparse. The founding example of such a phage, φVP882, detects a chemical communication signal molecule called DPO that is produced by diverse bacterial species. We investigated whether a family of VP882-like phages might exist that detect and respond to DPO. We find that indeed, VP882-like phages reside in DPO-producing bacterial species isolated at different times and geographic locations, suggesting their wide circulation in the environment. This discovery strengthens the evidence for the generality of phage-bacterial inter-domain chemical communication. 

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5. Development of an autonomous and bifunctional quorum-sensing circuit for metabolic flux control in engineered Escherichia coli

   https://doi.org/10.1073/pnas.1911144116  

   Dinh, Christina V.; Prather, Kristala L. (December 2019, Proceedings of the National Academy of Sciences)

   Metabolic engineering seeks to reprogram microbial cells to efficiently and sustainably produce value-added compounds. Since chemical production can be at odds with the cell’s natural objectives, strategies have been developed to balance conflicting goals. For example, dynamic regulation modulates gene expression to favor biomass and metabolite accumulation at low cell densities before diverting key metabolic fluxes toward product formation. To trigger changes in gene expression in a pathway-independent manner without the need for exogenous inducers, researchers have coupled gene expression to quorum-sensing (QS) circuits, which regulate transcription based on cell density. While effective, studies thus far have been limited to one control point. More challenging pathways may require layered dynamic regulation strategies, motivating the development of a generalizable tool for regulating multiple sets of genes. We have developed a QS-based regulation tool that combines components of the lux and esa QS systems to simultaneously and dynamically up- and down-regulate expression of 2 sets of genes. Characterization of the circuit revealed that varying the expression level of 2 QS components leads to predictable changes in switching dynamics and that using components from 2 QS systems allows for independent tuning capability. We applied the regulation tool to successfully address challenges in both the naringenin and salicylic acid synthesis pathways. Through these case studies, we confirmed the benefit of having multiple control points, predictable tuning capabilities, and independently tunable regulation modules. 

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