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Pseudomonas aeruginosa AmpR Is a Global Transcriptional Factor That Regulates Expression of AmpC and PoxB ß-Lactamases, Proteases, Quorum Sensing, and Other Virulence Factors

Kok-Fai Kong,¹ Suriya Ravi Jayawardena,¹ Shalaka Dayaram Indulkar,¹ Aimee del Puerto,¹ Chong-Lek Koh,^2, Niels Høiby,³ and Kalai Mathee^1*

Department of Biological Sciences, Florida International University, Miami, Florida 33199,¹ Institute of Biological Sciences (Genetics), University of Malaya, 50603 Kuala Lumpur, Malaysia,² Department of Clinical Microbiology, University Hospital of Copenhagen, Copenhagen, Denmark³

Received 21 February 2005/ Returned for modification 21 April 2005/ Accepted 25 August 2005

In members of the family Enterobacteriaceae, ampC, which encodes a ß-lactamase, is regulated by an upstream, divergently transcribed gene, ampR. However, in Pseudomonas aeruginosa, the regulation of ampC is not understood. In this study, we compared the characteristics of a P. aeruginosa ampR mutant, PAOampR, with that of an isogenic ampR⁺ parent. The ampR mutation greatly altered AmpC production. In the absence of antibiotic, PAOampR expressed increased basal ß-lactamase levels. However, this increase was not followed by a concomitant increase in the P_ampC promoter activity. The discrepancy in protein and transcription analyses led us to discover the presence of another chromosomal AmpR-regulated ß-lactamase, PoxB. We found that the expression of P. aeruginosa ampR greatly altered the ß-lactamase production from ampC and poxB in Escherichia coli: it up-regulated AmpC but down-regulated PoxB activities. In addition, the constitutive P_ampR promoter activity in PAOampR indicated that AmpR did not autoregulate in the absence or presence of inducers. We further demonstrated that AmpR is a global regulator because the strain carrying the ampR mutation produced higher levels of pyocyanin and LasA protease and lower levels of LasB elastase than the wild-type strain. The increase in LasA levels was positively correlated with the P_lasA, P_lasI, and P_lasR expression. The reduction in the LasB activity was positively correlated with the P_rhlR expression. Thus, AmpR plays a dual role, positively regulating the ampC, lasB, and rhlR expression levels and negatively regulating the poxB, lasA, lasI, and lasR expression levels.

Present address: National Institute of Education, Nanyang Technological University, Singapore 637616.

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