Characterization of a small PlcR-regulated gene co-expressed with cereolysin O

In the genomes of bacteria belonging to the B. cereus group, including Bacillus anthracis and Bacillus thuringiensis, a small gene encoding a 26-amino acid peptide was present in multicopy. One copy was always found upstream from the gene encoding Clo. In B. cereus ATCC 14579, the small gene and the clo gene are co-transcribed. Transcriptional fusions showed that the three paralogues identified in this strain were expressed in a PlcR-dependent manner. We propose to name these peptides Spp for small PlcR-regulated peptides. We show that a synthetic peptide corresponding to the deduced product of the spp genes displayed antibacterial activity.The co-expression of spp, a small PlcR-regulated multicopy gene with clo suggests a yet unidentified relationship between Spp and the cholesterol-dependent cytolysin in bacteria belonging to the B.cereus group.Bacillus cereus is an opportunistic pathogen of humans, causing local and systemic infections, and is a frequent cause of food poisoning. This species belongs to the B. cereus group, which includes the closely related species Bacillus anthracis, Bacillus thuringiensis, Bacillus weihenstephanensis,Bacillus mycoides and Bacillus pseudomycoides [1,2]. B. cereus produces several secreted proteins, including enterotoxins, cytolysins, phospholipases and proteases that may contribute to B. cereus pathogenicity. The expression of most of these virulence factors is controlled by the pleiotropic transcriptional activator PlcR [3,4]. This global regulator has been shown to contribute to B. cereus virulence in mice and insects [5] and in rabbit endophthalmitis [6]. Expression of the PlcR regulon is activated at the onset of the stationary phase of growth [7]. This activation results from cell-cell communication under the control of PapR, a small peptide that is exported, processed, and re-imported into bacterial cells in its mature form, presumably a pentapeptide, by the oligopeptide permease [8,9].Haemolysins of the cholesterol-depend