The CodY-dependent clhAB2 operon is involved in cell shape, chaining and autolysis in Bacillus cereus ATCC 14579

The Gram-positive pathogen Bacillus cereus is able to grow in chains of rod-shaped cells, but the regulation of chaining remains largely unknown. Here, we observe that glucose-grown cells of B . cereus ATCC 14579 form longer chains than those grown in the absence of glucose during the late exponential and transition growth phases, and identify that the clhAB 2 operon is required for this chain lengthening phenotype. The clhAB 2 operon is specific to the B . cereus group (i.e., B . thuringiensis , B . anthracis and B . cereus ) and encodes two membrane proteins of unknown function, which are homologous to the Staphylococcus aureus CidA and CidB proteins involved in cell death control within glucose-grown cells. A deletion mutant (Δ clhAB 2 ) was constructed and our quantitative image analyses show that Δ clhAB 2 cells formed abnormal short chains regardless of the presence of glucose. We also found that glucose-grown cells of Δ clhAB 2 were significantly wider than wild-type cells (1.47 μm ±CI 95% 0.04 vs 1.19 μm ±CI 95% 0.03, respectively), suggesting an alteration of the bacterial cell wall. Remarkably, Δ clhAB 2 cells showed accelerated autolysis under autolysis-inducing conditions, compared to wild-type cells. Overall, our data suggest that the B . cereus clhAB 2 operon modulates peptidoglycan hydrolase activity, which is required for proper cell shape and chain length during cell growth, and down-regulates autolysin activity. Lastly, we studied the transcription of clhAB 2 using a lacZ transcriptional reporter in wild-type, ccpA and codY deletion-mutant strains. We found that the global transcriptional regulatory protein CodY is required for the basal level of clhAB 2 expression under all conditions tested, including the transition growth phase while CcpA, the major global carbon regulator, is needed for the high-level expression of clhAB 2 in glucose-grown cells.