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Journal of Bacteriology, January 2004, p. 316-325, Vol. 186, No. 2
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.2.316-325.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

The Alternative Sigma Factor {sigma}B of Bacillus cereus: Response to Stress and Role in Heat Adaptation

Willem van Schaik,1,2 Marcel H. Tempelaars,1,2 Jeroen A. Wouters,1,2,{dagger} Willem M. de Vos,1 and Tjakko Abee1,2*

Wageningen Centre for Food Sciences,1 Laboratory of Food Microbiology, Wageningen University, Wageningen, The Netherlands2

Received 22 July 2003/ Accepted 8 October 2003

A gene cluster encoding the alternative sigma factor {sigma}B, three predicted regulators of {sigma}B (RsbV, RsbW, and RsbY), and one protein whose function is not known (Orf4) was identified in the genome sequence of the food pathogen Bacillus cereus ATCC 14579. Western blotting with polyclonal antibodies raised against {sigma}B revealed that there was 20.1-fold activation of {sigma}B after a heat shock from 30 to 42°C. Osmotic upshock and ethanol exposure also upregulated {sigma}B, albeit less than a heat shock. When the intracellular ATP concentration was decreased by exposure to carbonyl cyanide m-chlorophenylhydrazone (CCCP), only limited increases in {sigma}B levels were observed, revealing that stress due to ATP depletion is not an important factor in {sigma}B activation in B. cereus. Analysis of transcription of the sigB operon by Northern blotting and primer extension revealed the presence of a {sigma}B-dependent promoter upstream of the first open reading frame (rsbV) of the sigB operon, indicating that transcription of sigB is autoregulated. A second {sigma}B-dependent promoter was identified upstream of the last open reading frame (orf4) of the sigB operon. Production of virulence factors and the nonhemolytic enterotoxin Nhe in a sigB null mutant was the same as in the parent strain. However, {sigma}B was found to play a role in the protective heat shock response of B. cereus. The sigB null mutant was less protected against the lethal temperature of 50°C by a preadaptation to 42°C than the parent strain was, resulting in a more-than-100-fold-reduced survival of the mutant after 40 min at 50°C.

* Corresponding author. Mailing address: Laboratory of Food Microbiology, Wageningen University, Bomenweg 2, 6703 HD Wageningen, The Netherlands. Phone: 31-317-484981. Fax: 31-317-484978. E-mail: tjakko.abee{at}wur.nl.

{dagger} Present address: Department Flavour, Nutrition and Ingredients, NIZO Food Research, 6710 BA Ede, The Netherlands.

Journal of Bacteriology, January 2004, p. 316-325, Vol. 186, No. 2
0021-9193/04/$08.00+0     DOI: 10.1128/JB.186.2.316-325.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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