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Journal of Bacteriology, August 2000, p. 4180-4187, Vol. 182, No. 15
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

Coactivation of the RpoS-Dependent proP P2 Promoter by Fis and Cyclic AMP Receptor Protein

Sarah M. McLeod,1 Jimin Xu,1,dagger and Reid C. Johnson1,2,*

Department of Biological Chemistry, UCLA School of Medicine, Los Angeles, California 90095-1737,1 and Molecular Biology Institute, University of California, Los Angeles, Los Angeles, California 900952

Received 27 January 2000/Accepted 26 April 2000

The Escherichia coli proP P2 promoter, which directs the expression of an integral membrane transporter of proline, glycine betaine, and other osmoprotecting compounds, is induced upon entry into stationary phase to protect cells from osmotic shock. Transcription from the P2 promoter is completely dependent on RpoS (sigma 38) and Fis. Fis activates transcription by binding to a site centered at -41, which overlaps the promoter, where it makes a specific contact with the C-terminal domain of the alpha  subunit of RNA polymerase (alpha -CTD). We show here that Fis and cyclic AMP (cAMP) receptor protein (CRP)-cAMP collaborate to activate transcription synergistically in vitro. Coactivation both in vivo and in vitro is dependent on CRP binding to a site centered at -121.5, but CRP without Fis provides little activation. The contribution by CRP requires the correct helical phasing of the CRP site and a functional activation region 1 on CRP. We provide evidence that coactivation is achieved by Fis and CRP independently contacting each of the two alpha -CTDs. Efficient transcription in vitro requires that both activators must be preincubated with the DNA prior to addition of RNA polymerase.

* Corresponding author. Mailing address: Department of Biological Chemistry, UCLA School of Medicine, 10833 Le Conte Ave., Los Angeles, CA 90095-1737. Phone: (310) 825-7800. Fax: (310) 206-5272. E-mail: rcjohnson{at}mednet.ucla.edu.

dagger Present address: West Los Angeles VA Medical Center, Los Angeles, CA 90073.

Journal of Bacteriology, August 2000, p. 4180-4187, Vol. 182, No. 15
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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