Characterization of the Operon Encoding the Alternative ς B Factor from Bacillus anthracis and Its Role in Virulence

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Abstract
The operon encoding the general stress transcription factor ς B and two proteins of its regulatory network, RsbV and RsbW, was cloned from the gram-positive bacterium Bacillus anthracis by PCR amplification of chromosomal DNA with degenerate primers, by inverse PCR, and by direct cloning. The gene cluster was very similar to the Bacillus subtilis sigB operon both in the primary sequences of the gene products and in the order of its three genes. However, the deduced products of sequences upstream and downstream from this operon showed no similarity to other proteins encoded by the B. subtilis sigB operon. Therefore, the B. anthracis sigB operon contains three genes rather than eight as in B. subtilis . The B. anthracis operon is preceded by a ς B -like promoter sequence, the expression of which depends on an intact ς B transcription factor in B. subtilis . It is followed by another open reading frame that is also preceded by a promoter sequence similarly dependent on B. subtilis ς B . We found that in B. anthracis , both these promoters were induced during the stationary phase and induction required an intact sigB gene. The sigB operon was induced by heat shock. Mutants from which sigB was deleted were constructed in a toxinogenic and a plasmidless strain. These mutants differed from the parental strains in terms of morphology. The toxinogenic sigB mutant strain was also less virulent than the parental strain in the mouse model. B. anthracis ς B may therefore be a minor virulence factor.