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Journal of Bacteriology, October 2009, p. 6211-6218, Vol. 191, No. 20
0021-9193/09/$08.00+0     doi:10.1128/JB.00888-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

The Multifaceted Proteins MvaT and MvaU, Members of the H-NS Family, Control Arginine Metabolism, Pyocyanin Synthesis, and Prophage Activation in Pseudomonas aeruginosa PAO1{triangledown}

Congran Li,1,2 Hassan Wally,1 Shearman J. Miller,1 and Chung-Dar Lu1*

Department of Biology, Georgia State University, Atlanta, Georgia 30303,1 Laboratory of Pharmacology, Institute of Medicinal Biotechnology, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China2

Received 8 July 2009/ Accepted 4 August 2009

The MvaT and MvaU proteins belonging to the H-NS family were identified as DNA-binding proteins that interact with the regulatory region of the aotJQMOP-argR operon for arginine uptake and regulation. Recombinant MvaT and MvaU proteins were purified, and binding of these purified proteins to the aotJ regulatory region was demonstrated using electromobility shift assays. Polyclonal antibodies against purified MvaT and MvaU were prepared and employed in supershift assays to support these observations. Knockout mutations resulting in a single lesion in mvaT or mvaU, as well as knockout mutations resulting in double lesions, were constructed using biparental conjugation, and the absence of MvaT and MvaU in the resulting mutants was confirmed by immunoblot analysis. Using measurements of the β-galactosidase activities from aotJ::lacZ fusions in the mutants and the parental strain, it was found that MvaT and MvaU serve as repressors in control of aotJ expression. The effects of MvaT and MvaU on pyocyanin synthesis and CupA fimbrial expression in these mutants were also analyzed. Pyocyanin synthesis was induced in the single mutants but was completely abolished in the double mutant, suggesting that there is a complicated regulatory scheme in which MvaT and MvaU are essential elements. In comparison, MvaT had a more profound role than MvaU as a repressor of cupA expression; however, a combination of MvaT depletion and MvaU depletion had a strong synergistic effect on cupA. Moreover, prophage Pf4 integrated into the chromosome of Pseudomonas aeruginosa PAO1 was activated in an mvaT mvaU double mutant but not in a single mutant. These results were supported by purification and nucleotide sequencing of replicative-form DNA and by the release of phage particles in plaque assays. In summary, the mvaT mvaU double mutant was viable, and depletion of MvaT and MvaU had serious effects on a variety of physiological functions in P. aeruginosa.

* Corresponding author. Mailing address: Department of Biology, Georgia State University, 24 Peachtree Center Avenue, Atlanta, GA 30303. Phone: (404) 413-5395. Fax: (404) 413-5301. E-mail: biocdl{at}langate.gsu.edu

{triangledown} Published ahead of print on 14 August 2009.

Journal of Bacteriology, October 2009, p. 6211-6218, Vol. 191, No. 20
0021-9193/09/$08.00+0     doi:10.1128/JB.00888-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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