Regulation and function of Pseudomonas drug efflux pumps

Summary

Principal Investigator: Herbert Schweizer
Abstract: DESCRIPTION (provided by applicant): Pseudomonas aeruginosa is an opportunistic pathogen that causes a multitude of infections, which are difficult to treat because of this bacterium's intrinsic and acquired antibiotic resistance. This antibiotic resistance can be attributed to synergy between a low-permeability outer membrane (OM) and active efflux from the cell. The genome of P. aeruginosa encodes 35 proposed drug efflux systems belonging to five different families, but our working hypothesis is that the clinically relevant efflux pumps contributing to intrinsic and/or acquired drug resistance belong to the resistance nodulation division (RND) family which contains 12 members. Genome sequence and expression analysis of these systems poses several questions that remain unanswered. First, because only a subset of the RND operons contains genes for OM channel proteins, the question is whether these systems must recruit other OM channels from elsewhere on the genome to function as tripartite efflux systems for drug efflux across the entire cell envelope, as current dogma says, or whether they can function as two-component systems for other substrates. Second, with the exception of a single pump, the other RND pumps are tightly regulated but the regulatory networks governing their expression and the inducing substrates remain unknown. Our other working hypotheses therefore are: (i) efflux pumps not encoding their own OM channels may either recruit hitherto unidentified proteins for function with certain substrates but may also function as two-component efflux systems for other substrates; (ii) Efflux pump expression is governed by specific and probably also global transcriptional regulators. We will test these hypotheses using the recently discovered MexJK efflux pump that is normally silent and does not encode its own OM channel. We also propose to amend these studies to probe whether any global transcriptional regulators may be involved in RND efflux pump expression. Specifically we propose to: Aim 1-Establish the regulation of MexJK by its cognate regulator MexL using biochemical and molecular studies; identify an inducer that may be present in certain growth media; identify a putative activating factor present in P. aeruginosa but absent in E. coil. Aim 2: Establish the molecular architecture of MexJK, specifically its OM membrane channel requirement, if any, using genetic and biochemical methods. Aim 3: Probe other, perhaps global regulators of efflux operon expression, specifically MexS and PA4878.
Funding Period: 2003-05-01 - 2007-04-30
more information: NIH RePORT

Top Publications

  1. ncbi Bacterial resistance to antibiotics: active efflux and reduced uptake
    Ayush Kumar
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO, USA
    Adv Drug Deliv Rev 57:1486-513. 2005
  2. ncbi Understanding efflux in Gram-negative bacteria: opportunities for drug discovery
    Herbert P Schweizer
    Colorado State University, IDRC at Foothills Campus, Department of Microbiology, Immunology and Pathology, Fort Collins, CO 80523 0922, USA
    Expert Opin Drug Discov 7:633-42. 2012
  3. ncbi Global transcriptional responses to triclosan exposure in Pseudomonas aeruginosa
    Rungtip Chuanchuen
    Department of Veterinary Public Health, Faculty of Veterinary Science, Chulalongkorn University, Pathumwan, Bangkok 10330, Thailand
    Int J Antimicrob Agents 40:114-22. 2012
  4. pmc Evidence of MexT-independent overexpression of MexEF-OprN multidrug efflux pump of Pseudomonas aeruginosa in presence of metabolic stress
    Ayush Kumar
    Antimicrobial Resistance Research Group, Applied Bioscience Program, Faculty of Health Sciences, University of Ontario Institute of Technology, Oshawa, Ontario, Canada
    PLoS ONE 6:e26520. 2011
  5. pmc Molecular characterization of MexL, the transcriptional repressor of the mexJK multidrug efflux operon in Pseudomonas aeruginosa
    Rungtip Chuanchuen
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523 1682, USA
    Antimicrob Agents Chemother 49:1844-51. 2005
  6. pmc Substrate-dependent utilization of OprM or OpmH by the Pseudomonas aeruginosa MexJK efflux pump
    Rungtip Chuanchuen
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, USA
    Antimicrob Agents Chemother 49:2133-6. 2005
  7. pmc Molecular basis of azithromycin-resistant Pseudomonas aeruginosa biofilms
    Richard J Gillis
    Department of Microbiology and Immunology, Box 672, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA
    Antimicrob Agents Chemother 49:3858-67. 2005
  8. pmc Identification and characterization of TriABC-OpmH, a triclosan efflux pump of Pseudomonas aeruginosa requiring two membrane fusion proteins
    Takehiko Mima
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523 1682, USA
    J Bacteriol 189:7600-9. 2007

Scientific Experts

  • Herbert Schweizer
  • Rungtip Chuanchuen
  • Ayush Kumar
  • Takehiko Mima
  • Richard J Gillis
  • Margarita Gomez-Escalada
  • Swati Joshi
  • Kimberly G White
  • Naomasa Gotoh
  • Victoria E Wagner
  • Jared B Gaynor
  • Barbara H Iglewski
  • Kyoung Hee Choi
  • Takeshi Murata
  • RoxAnn Karkhoff-Schweizer

Detail Information

Publications8

  1. ncbi Bacterial resistance to antibiotics: active efflux and reduced uptake
    Ayush Kumar
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO, USA
    Adv Drug Deliv Rev 57:1486-513. 2005
    ..Recent advances in understanding the physical structures, function and regulation of efflux systems will facilitate exploitation of pumps as new drug targets...
  2. ncbi Understanding efflux in Gram-negative bacteria: opportunities for drug discovery
    Herbert P Schweizer
    Colorado State University, IDRC at Foothills Campus, Department of Microbiology, Immunology and Pathology, Fort Collins, CO 80523 0922, USA
    Expert Opin Drug Discov 7:633-42. 2012
    ..However, recent advances in our understanding of efflux in these bacteria provide opportunities and assets for drug discovery...
  3. ncbi Global transcriptional responses to triclosan exposure in Pseudomonas aeruginosa
    Rungtip Chuanchuen
    Department of Veterinary Public Health, Faculty of Veterinary Science, Chulalongkorn University, Pathumwan, Bangkok 10330, Thailand
    Int J Antimicrob Agents 40:114-22. 2012
    ..The profound perturbations of cellular metabolism via specific and global mechanisms may explain why triclosan is such a potent antimicrobial in susceptible bacteria...
  4. pmc Evidence of MexT-independent overexpression of MexEF-OprN multidrug efflux pump of Pseudomonas aeruginosa in presence of metabolic stress
    Ayush Kumar
    Antimicrobial Resistance Research Group, Applied Bioscience Program, Faculty of Health Sciences, University of Ontario Institute of Technology, Oshawa, Ontario, Canada
    PLoS ONE 6:e26520. 2011
    ..Regulation of mexEF-oprN operon expression is multifaceted with the MexT activator being one of the most prominent regulatory proteins...
  5. pmc Molecular characterization of MexL, the transcriptional repressor of the mexJK multidrug efflux operon in Pseudomonas aeruginosa
    Rungtip Chuanchuen
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523 1682, USA
    Antimicrob Agents Chemother 49:1844-51. 2005
    ..The MexL-protected region contains two inverted GTATTT repeats, and their location in the protected region and overlap with the mexL and mexJ promoter sequences strongly support a role in MexL binding...
  6. pmc Substrate-dependent utilization of OprM or OpmH by the Pseudomonas aeruginosa MexJK efflux pump
    Rungtip Chuanchuen
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523, USA
    Antimicrob Agents Chemother 49:2133-6. 2005
    ..This is the first report of natural utilization of multiple OMPs by a given resistance nodulation cell division transporter/membrane fusion protein pair...
  7. pmc Molecular basis of azithromycin-resistant Pseudomonas aeruginosa biofilms
    Richard J Gillis
    Department of Microbiology and Immunology, Box 672, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA
    Antimicrob Agents Chemother 49:3858-67. 2005
    ..mexA, which is constitutively expressed in planktonic cells, was uniformly expressed in biofilms regardless of the presence of AZM. These data indicate that the MexCD-OprJ pump acts as a biofilm-specific mechanism for AZM resistance...
  8. pmc Identification and characterization of TriABC-OpmH, a triclosan efflux pump of Pseudomonas aeruginosa requiring two membrane fusion proteins
    Takehiko Mima
    Department of Microbiology, Immunology and Pathology, Colorado State University, Fort Collins, CO 80523 1682, USA
    J Bacteriol 189:7600-9. 2007
    ..TriABC is the fifth RND pump in P. aeruginosa shown to efficiently efflux triclosan, supporting the notion that efflux is the primary mechanism responsible for this bacterium's high intrinsic and acquired triclosan resistance...