|Abstract or Summary
- Cell-to-cell communication by chemical signals, termed quorum sensing (QS), is a common regulatory scheme in the microbial world. Pseudomonas aeruginosa¸ an opportunistic pathogen of burn wounds and cystic fibrosis lungs, uses QS to control the expression of hundreds of genes, particularly those necessary for population level benefits such as biofilm formation and secretion of extracellular virulence factors (so-called public goods). P. aeruginosa has two QS systems, las and rhl, that use diffusible acyl-homoserine lactone signals (acyl-HSL). Each system is comprised of a signal synthase (LasI and RhlI) and a cognate receptor transcription factor (LasR and RhlR). Under certain conditions, the las system regulates the rhl system. The circuitry is subject to additional regulation as accumulation of signal is necessary, but not sufficient to activate most QS-controlled genes. From a social evolution perspective, P. aeruginosa QS is considered a cooperative behavior that can be exploited by lasR mutant cheaters that do not contribute public goods. Here, we answer two questions: how social cheating influences the evolution of quorum-sensing inhibitor (QSI) resistance, and what nutritional cues promote QS gene expression. We designed a proof-of-concept experiment to understand how bacterial social interactions affect the evolution of resistance to QSI antivirulence. We cultured QS-deficient mutants with small proportions of QS-proficient wild-type to mimic QSI-sensitive and QSI-resistant cells, respectively. We employed two different carbon sources that are degraded by QS-controlled extracellular, secreted (public) or cell-associated (private) enzymes. We found that QSI-sensitive mimics (QS-deficient cells) behave as social cheaters that delayed population growth and prevented enrichment of QSI-resistant mimics (QS-proficient cells) only when nutrient acquisition was public, suggesting that QSI resistance would not spread. To answer the second question, we used minimal medium batch and chemostat cultures to demonstrate that specific macronutrient starvation coupled with growth rate reduction induces expression of secreted factors directly controlled by the las and rhl QS systems. The rhl system was more responsive to starvation and growth rate reduction as the transcriptional regulator RhlR and its cognate acyl-HSL were strongly induced. Our results also showed that a slow growth rate inverted the las-to-rhl acyl-HSL signal ratio, previously considered a distinguishing characteristic between planktonic and biofilm lifestyles. Importantly, expression level depended on the elemental composition of the secreted product and increased only when the limiting nutrient was not also a building block. Such supply-driven regulation is metabolically prudent as it reduces the costs associated with public goods production, which in turn can help limit the metabolic advantage of non-secreting social cheaters. Our results define the physiological basis for the co-regulation of QS-controlled genes by stress responses. They have implications for the evolutionary stability of microbial cooperation as well as for the efficacy of antivirulence drugs and the emergence of resistance to these drugs.