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Journal of Bacteriology, April 2000, p. 2077-2087, Vol. 182, No. 8
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.

RegA, Iron, and Growth Phase Regulate Expression of the Pseudomonas aeruginosa tol-oprL Gene Cluster

Kangmin Duan, Eric R. Lafontaine,dagger Sonali Majumdar, and Pamela A. Sokol*

Department of Microbiology and Infectious Diseases, University of Calgary Health Sciences Centre, Calgary, Alberta T2N 4N1, Canada

Received 19 August 1999/Accepted 12 January 2000

The tol-oprL region in Pseudomonas aeruginosa appears to be involved in pyocin uptake and required for cell viability. The complete nucleotide sequences of the tolQRA and oprL genes as well as the incomplete sequences of tolB and orf2 have been previously reported. In addition, the sequence of a P. aeruginosa iron-regulated gene (pig6) has been described and found to share homology with an open reading frame located upstream of the Escherichia coli tolQRA genes (U. A. Ochsner and M. L. Vasil, Proc. Natl. Acad. Sci. USA 93:4409-4414, 1996). In this study, we cloned the remainder of the P. aeruginosa tol-oprL gene cluster and determined its nucleotide sequence. This cluster was found to consist of seven genes in the order orf1 tolQ tolR tolA tolB oprL orf2. Transcriptional analysis of this gene cluster was performed by detecting the presence of mRNAs spanning adjacent genes as well as by using a promoterless lacZ reporter gene fused to each of the seven genes contained in the tol-oprL locus. The results show that there are three major transcriptional units or operons in this region, orf1-tolQRA, tolB, and oprL-orf2, in contrast to the E. coli tol-pal region, where there are only two operons, orf1-tolQRA and tolB-pal-orf2. Analysis of gene expression indicated that the tol-oprL genes of P. aeruginosa are both iron and growth phase modulated. The first operon, orf1-tolQRA, is iron regulated throughout growth, but iron-regulated expression of tolB and oprL fusions occurs only in late log phase. The expression of the three operons was significantly less repressed by iron in fur mutants than in the wild-type strain, suggesting the involvement of Fur in the iron regulation of all three operons. RegA is a positive yet nonessential regulator of tol-oprL expression.

* Corresponding author. Mailing address: Department of Microbiology and Infectious Diseases, University of Calgary Health Sciences Centre, 3330 Hospital Dr. NW, Calgary, Alberta, Canada T2N 4N1. Phone: (403) 220-6037. Fax: (403) 270-2772. E-mail: psokol{at}ucalgary.ca.

dagger Present address: Department of Microbiology, University of Texas Southwestern Medical Center, Dallas, Tex.

Journal of Bacteriology, April 2000, p. 2077-2087, Vol. 182, No. 8
0021-9193/00/$04.00+0
Copyright © 2000, American Society for Microbiology. All rights reserved.


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