# An anticipatory mechanism enhances the cooperative behaviors of quorum sensing mutants in Pseudomonas aeruginosa

## TL;DR

This study reveals how a regulatory mechanism involving PsdR and LasR228 stabilizes cooperation in Pseudomonas aeruginosa, preventing cheating and enhancing pathogenicity.

## Contribution

A novel anticipatory regulatory mechanism involving PsdR and LasR228 is identified to stabilize cooperation in microbial populations.

## Key findings

- Inactivation of PsdR restores function in LasR228 variant strains, enhancing cooperative behaviors.
- PsdR-null-LasR228 strains exhibit increased cooperation and pathogenicity compared to LasR-null strains.
- PsdR-mediated regulation serves as an anticipatory control against cheating LasR variant strains.

## Abstract

Social interactions substantially influence the dynamics and functions of microbial communities. Cooperative behaviors serve to benefit populations, yet they are often exploited by cheating cells, thus creating a conflict between individuals in the microbial population. However, the underlying mechanisms by which cooperative behaviors are stabilized are incompletely elucidated. Here, we used quorum sensing (QS) as a model of cooperation, and functionally studied QS regulator LasR variant strains in the context of cooperative behaviors. We found that a LasR228 variant strain, bearing a non-conserved substitution in LasR, exhibited minimal LasR-dependent phenotypes. However, the function of this LasR228 variant strain was restored by inactivation of the transcriptional repressor PsdR, and the phenotypes of this variant strain were similar to the parental strain. Furthermore, we illustrate a post-transcriptional regulatory mechanism responsible for the activation of the LasR228 variant. Unlike LasR228, the PsdR-null-LasR228 strain demonstrated cooperative behaviors in competition with the LasR-null strain. Since psdR mutations precede the emergence of LasR variants in the evolution of P. aeruginosa using casein broth, this PsdR-mediated cooperative mechanism serves as an anticipatory control against potential cheating LasR variant strains. Additionally, our cell-killing assay showed that the cooperative PsdR-null-LasR228 strain was associated with increased bacterial pathogenicity to eukaryotic host cells. In conclusion, our study reveals the functional plasticity of LasR variants, which can be modulated by secondary mutations, affecting cooperation and conflict within populations. Our identification of a novel cooperative molecular mechanism offers insight into the maintenance of cooperation within microbial communities.

This research uncovers a new mechanism that stabilizes cooperation in microbial populations by studying quorum sensing (QS) in Pseudomonas aeruginosa. Our findings identified a regulatory pathway where the PsdR protein modulates the function of a QS regulator variant (LasR228), influencing cooperative behaviors and pathogenicity. By inactivating PsdR, the variant’s cooperative behaviors were enhanced, facilitating the production of public goods. This finding highlights how microbial communities can adapt through secondary mutations to preserve cooperative dynamics, which are essential for their survival and pathogenic traits. Understanding these mechanisms provides valuable insights into microbial evolution and the potential development of therapies targeting microbial cooperation to mitigate infections.