the addition of autoinducers restored CRISPR–Cas immunity in the double mutant
In P. aeruginosa, adaptive immunity is mediated, in part, by the helicase and nuclease Cas3. In accordance with the hypothesis that quorum sensing activates immunity, the expression of cas3 increased at high cell density. Quorum sensing depends on signalling molecules, known as autoinducers, and their receptors, and the two main quorum sensing systems in P. aeruginosa are LasI–LasR and RhlI–RhlR. When both autoinducer synthases, lasI and rhlI, were deleted, the expression of cas3 decreased at high cell densities; however, expression could be rescued by the addition of the autoinducers. Furthermore, when the double mutant was transformed with a plasmid that contained a protospacer sequence that is targeted by the CRISPR–Cas system, more mutant bacteria retained the plasmid than wild-type bacteria, which eliminated the plasmid after several hours, when quorum sensing was strongly induced. Again, the addition of autoinducers restored CRISPR–Cas immunity in the double mutant, which confirmed that quorum sensing is required for optimal CRISPR–Cas activity. Finally, the authors also showed that the double mutant acquired new CRISPR spacers only half as often as wild-type bacteria.
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