CRISPR-Cas are prokaryotic adaptive immune systems that provide protection against bacteriophage (phage) and other parasites. Little is known about how CRISPR-Cas systems are regulated, preventing prediction of phage dynamics in nature and manipulation of phage resistance in clinical settings. Here, we show that the bacterium Pseudomonas aeruginosa PA14 uses the cell-cell communication process, called quorum sensing, to activate cas gene expression, to increase CRISPR-Cas targeting of foreign DNA, and to promote CRISPR adaptation, all at high cell density. This regulatory mechanism ensures maximum CRISPR-Cas function when bacterial populations are at highest risk for phage infection. We demonstrate that CRISPR-Cas activity and acquisition of resistance can be modulated by administration of proand antiquorum-sensing compounds. We propose that quorum-sensing inhibitors could be used to suppress the CRISPR-Cas adaptive immune system to enhance medical applications, including phage therapies.quorum sensing | CRISPR | immunity | phage | phage defense

Quorum sensing is a process of cell-cell communication that bacteria use to regulate collective behaviors. Quorum sensing depends on the production, detection, and group-wide response to extracellular signal molecules called autoinducers. In many bacterial species, quorum sensing controls virulence factor production. Thus, disrupting quorum sensing is considered a promising strategy to combat bacterial pathogenicity. Several members of a family of naturally produced plant metabolites called flavonoids inhibit biofilm formation by an unknown mechanism. Here, we explore this family of molecules further, and we demonstrate that flavonoids specifically inhibit quorum sensing via antagonism of the autoinducer-binding receptors, LasR and RhlR. Structure-activity relationship analyses demonstrate that the presence of two hydroxyl moieties in the flavone A-ring backbone are essential for potent inhibition of LasR/RhlR. Biochemical analyses reveal that the flavonoids function non-competitively to prevent LasR/RhlR DNA binding. Administration of the flavonoids to alters transcription of quorum sensing-controlled target promoters and suppresses virulence factor production, confirming their potential as anti-infectives that do not function by traditional bacteriocidal or bacteriostatic mechanisms.

Bacteria use a cell-cell communication process called quorum sensing (QS) to orchestrate collective behaviors. QS relies on the group-wide detection of molecules called autoinducers (AI).