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J Bacteriol, February 1998, p. 785-792, Vol. 180, No. 4
0021-9193/98/$04.00+0
Copyright © 1998, American Society for Microbiology. All rights reserved.

Constricted Flux through the Branched-Chain Amino Acid Biosynthetic Enzyme Acetolactate Synthase Triggers Elevated Expression of Genes Regulated by rpoS and Internal Acidification

Tina K. Van Dyk,1,2,* Brenda L. Ayers,1,dagger Robin W. Morgan,2 and Robert A. Larossa1

Central Research and Development Department, DuPont Co., Wilmington, Delaware 19880-0173,1 and Department of Animal and Food Sciences, University of Delaware, Newark, Delaware 19717-13032

Received 22 September 1997/Accepted 11 December 1997

The first common enzyme of isoleucine and valine biosynthesis, acetolactate synthase (ALS), is specifically inhibited by the herbicide sulfometuron methyl (SM). To further understand the physiological consequences of flux alterations at this point in metabolism, Escherichia coli genes whose expression was induced by partial inhibition of ALS were sought. Plasmid-based fusions of random E. coli DNA fragments to Photorhabdus luminescens luxCDABE were screened for bioluminescent increases in actively growing liquid cultures slowed 25% by the addition of SM. From more than 8,000 transformants, 12 unique SM-inducible promoter-lux fusions were identified. The lux reporter genes were joined to seven uncharacterized open reading frames, f253a, f415, frvX, o513, o521, yciG, and yohF, and five known genes, inaA, ldcC, osmY, poxB, and sohA. Inactivation of the rpoS-encoded sigma factor, sigma S, reduced basal expression levels of six of these fusions 10- to 200-fold. These six genes defined four new members of the sigma S regulon, f253a, ldcC, yciG, and yohF, and included two known members, osmY and poxB. Furthermore, the weak acid salicylate, which causes cytoplasmic acidification, also induced increased bioluminescence from seven SM-inducible promoter-lux fusion-containing strains, namely, those with fusions of the sigma S-controlled genes and inaA. The pattern of gene expression changes suggested that restricted ALS activity may result in intracellular acidification and induction of the sigma S-dependent stress response.

* Corresponding author. Mailing address: Central Research and Development Department, DuPont Co., P.O. Box 80173, Wilmington, DE 19880-0173. Phone: (302) 695-1430. Fax: (302) 695-9183. E-mail: Tina.K.Van-Dyk{at}usa.dupont.com.

dagger Present address: Rockefeller University, New York, NY 10021.




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