Sensing danger.

CRISPR-Cas loci encode for an adaptive immune system in prokaryotes that provides defense against viruses (1) and plasmids (2) that infect these organisms. CRISPR loci contain a series of repetitive sequences intercalated with short sequences derived from invading viruses and plasmids (3–5). These short sequences, called spacers, are acquired from the invader upon infection during an “immunization” event (1). This is the first phase of CRISPR-Cas immunity. The information stored in spacers is then used to target the invader in the second phase of the CRISPR immune response (6). Each spacer sequence is transcribed and processed as a short RNA called CRISPR RNA (crRNA) that directs RNA-guided Cas nucleases to their matching target in the invader’s genome to cleave it. Little is known about the regulation of the CRISPR-Cas immune response. The CRISPR-Cas locus present in Escherichia coli has been shown to be tightly repressed by HNS, a negative regulator of widespread effects in bacteria (7). Repression can be relieved by the LeuO activator (8), but environmental or physiological conditions that lead to the expression of the CRISPR-Cas machinery have not been identified. In PNAS, Høyland-Kroghsbo et al. (9) describe the regulation of the CRISPR-Cas locus of the bacterial pathogen Pseudomonas aeruginosa through quorum sensing (QS) pathways. QS is a mechanism that bacteria use to communicate with each other and organize collective behaviors (10), mediated by genes that are responsible for the production and detection of extracellular chemicals known as autoinducers. Upon bacterial growth, the increase in the concentration of autoinducers triggers a signaling cascade that modifies the response of the bacterial population to different environmental cues (Fig. 1). Høyland-Kroghsbo et al. (9) first investigate whether the expression of the cas (CRISPR-associated) genes is affected by QS. cas genes flank the CRISPR array of repeats and spacers and are responsible for the execution of both phases of the CRISPR-Cas immune response (6). It was found that cas transcription is significantly increased at high cell density, and that this increase is dependent on both lasI and rhlI, the genes encoding for QS autoinducer synthases. Høyland-Kroghsbo et al. (9) then check for the effects of the deletion of these genes in both stages of the CRISPR-Cas immune response: targeting and immunization. The expectation is that the down-regulation of casgenes that results from the absence of autoinducer production in the lasI,rhlI double mutant should negatively affect CRISPR-Cas immunity. Targeting is tested through transformation with a plasmid harboring a target sequence, which is considerably Fig. 1. Regulation of CRISPR-Cas activity by QS. At low cell density, low autoinducer (pink dot) concentrations result in the repression of CRISPR-cas loci through the QS pathway, keeping the CRISPR-Cas immune response at a minimum. In contrast, at high densities of bacteria (when they are most susceptible to phage attack) the increase in the concentration of autoinducer triggers the de-repression of CRISPR-cas loci. Expression of the Cas protein complex and crRNA guide prepares the cell for the cleavage (arrowhead) of incoming viruses.