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Applied and Environmental Microbiology, August 2004, p. 4941-4949, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4941-4949.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Differential Gene Expression To Investigate the Effect of (5Z)-4-Bromo- 5-(Bromomethylene)-3-Butyl-2(5H)-Furanone on Bacillus subtilis

Dacheng Ren,1,{dagger} Laura A. Bedzyk,2 Peter Setlow,3 Dacre F. England,4 Staffan Kjelleberg,4 Stuart M. Thomas,2 Rick W. Ye,2 and Thomas K. Wood1*

Departments of Chemical Engineering and Molecular & Cell Biology, University of Connecticut, Storrs, Connecticut 06269-3222,1 Experimental Station E328/B33, DuPont Central Research and Development, Wilmington, Delaware 19880,2 Department of Biochemistry, University of Connecticut Health Center, Farmington, Connecticut 06032,3 School of Biotechnology and Biomolecular Sciences and Centre for Marine Biofouling and Bio-Innovation, University of New South Wales, Sydney, NSW 2052, Australia4

Received 19 January 2004/ Accepted 16 March 2004

(5Z)-4-Bromo-5-(bromomethylene)-3-butyl-2(5H)-furanone (furanone) from the red marine alga Delisea pulchra was found previously to inhibit the growth, swarming, and biofilm formation of gram-positive bacteria. Using the gram-positive bacterium Bacillus subtilis as a test organism, we observed cell killing by 20 µg of furanone per ml, while 5 µg of furanone per ml inhibited growth approximately twofold without killing the cells. To discover the mechanism of this inhibition on a genetic level and to investigate furanone as a novel antibiotic, full-genome DNA microarrays were used to analyze the gene expression profiles of B. subtilis grown with and without 5 µg of furanone per ml. This agent induced 92 genes more than fivefold (P < 0.05) and repressed 15 genes more than fivefold (P < 0.05). The induced genes include genes involved in stress responses (such as the class III heat shock genes clpC, clpE, and ctsR and the class I heat shock genes groES, but no class II or IV heat shock genes), fatty acid biosynthesis, lichenan degradation, transport, and metabolism, as well as 59 genes with unknown functions. The microarray results for four genes were confirmed by RNA dot blotting. Mutation of a stress response gene, clpC, caused B. subtilis to be much more sensitive to 5 µg of furanone per ml (there was no growth in 8 h, while the wild-type strain grew to the stationary phase in 8 h) and confirmed the importance of the induction of this gene as identified by the microarray analysis.

* Corresponding author. Mailing address: Departments of Chemical Engineering and Molecular & Cell Biology, University of Connecticut, 191 Auditorium Rd., U-3222, Storrs, CT 06269-3222. Phone: (860) 486-2483. Fax: (860) 486-2959. E-mail: twood{at}engr.uconn.edu.

{dagger} Present address: School of Chemical and Biomolecular Engineering, Cornell University, Ithaca, NY 14853.

Applied and Environmental Microbiology, August 2004, p. 4941-4949, Vol. 70, No. 8
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.8.4941-4949.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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