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Journal of Bacteriology, April 1999, p. 2110-2117, Vol. 181, No. 7
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.

Binding Site Recognition by Rns, a Virulence Regulator in the AraC Family

George P. Munson and June R. Scott*

Department of Microbiology and Immunology, Emory University Health Sciences Center, Atlanta, Georgia 30322

Received 28 October 1998/Accepted 15 January 1999

The expression of CS1 pili by enterotoxigenic strains of Escherichia coli is regulated at the transcriptional level and requires the virulence regulator Rns, a member of the AraC family of regulatory proteins. Rns binds at two separate sites upstream of Pcoo (the promoter of CS1 pilin genes), which were identified in vitro with an MBP::Rns fusion protein in gel mobility and DNase I footprinting assays. At each site, Rns recognizes asymmetric nucleotide sequences in two regions of the major groove and binds along one face of the DNA helix. Both binding sites are required for activation of Pcoo in vivo, because mutagenesis of either site significantly reduced the level of expression from this promoter. Thus, Rns regulates the expression of CS1 pilin genes directly, not via a regulatory cascade. Analysis of Rns-nucleotide interactions at each site suggests that binding sites for Rns and related virulence regulators are not easily identified because they do not bind palindromic or repeated sequences. A strategy to identify asymmetric binding sites is presented and applied to locate potential binding sites upstream of other genes that Rns can activate, including those encoding the CS2 and CFA/I pili of enterotoxigenic E. coli and the global regulator virB of Shigella flexneri.

* Corresponding author. Mailing address: Department of Microbiology and Immunology, Emory University Health Sciences Center, Atlanta, GA 30322. Phone: (404) 727-0402. Fax: (404) 727-8999. E-mail: Scott{at}microbio.emory.edu.

Journal of Bacteriology, April 1999, p. 2110-2117, Vol. 181, No. 7
0021-9193/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.


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