The dynamic response of quorum-sensing to density is robust to signal supplementation and signal synthase knockouts

Abstract

Quorum sensing (QS) is a widespread mechanism of environment sensing and behavioral coordination in bacteria. At its core, QS is based on the production, sensing and response to small signaling molecules. Previous work with Pseudomonas aeruginosa shows that QS can be used to achieve quantitative resolution and deliver a dosed response to the bacterias density environment, implying a sophisticated mechanism of control. To shed light on how the mechanistic signal components contribute to graded responses to density, we assess the impact of genetic (AHL signal synthase deletion) and/or signal supplementation (exogenous AHL and PQS addition) perturbations on lasB reaction-norms to changes in density. Our approach condenses data from 2,000 timeseries (over 74,000 individual observations) into a comprehensive view of QS-controlled gene expression across variation in genetic, environmental, and signal determinants of lasB expression. We first confirm that deleting either ({Delta}lasI, {Delta}rhlI) or both ({Delta}lasIrhlI) signal synthase genes attenuates QS response. In the {Delta}rhlI background we show persistent yet attenuated density-dependent lasB expression due to native 3-oxo-C12 signaling. We then test if density-independent quantities of signal (3-oxo-C12, C4, PQS or combined) added to the WT either flatten or increase the reaction norm and find that the WT response is robust to all tested concentrations of signal, alone or in combination. We then move to progressively supplementing the genetic knockouts and find that cognate signal supplementation ({Delta}lasI+3-oxo-C12, {Delta}rhlI+C4) is sufficient to restore lasB expression and as well as reactivity to density encoded by the intact signal synthase. We also find that dual supplementation of the double synthase knockout restores expression but does not flatten the reaction norm. Despite adding a density-independent amount of AHL, the double signal synthase can still quantitively sense density. Our results show that a positive reaction norm to density is robust to multiple combinations of gene deletion and density-independent signal supplementation and that while density-independent signal supplementation can increase mean expression, the WT QS still retains the ability to quantitatively respond to density. Our work develops a modular approach to query the robustness and mechanistic bases of the central environmental sensing phenotype of quorum sensing.