<i>Bordetella</i>AlcS Transporter Functions in Alcaligin Siderophore Export and Is Central to Inducer Sensing in Positive Regulation of Alcaligin System Gene Expression

  • Timothy J. Brickman
    Department of Microbiology, University of Minnesota Medical School, Minneapolis, Minnesota 55455-0312
  • Sandra K. Armstrong
    Department of Microbiology, University of Minnesota Medical School, Minneapolis, Minnesota 55455-0312

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<jats:title>ABSTRACT</jats:title><jats:p><jats:italic>Bordetella pertussis</jats:italic>and<jats:italic>Bordetella bronchiseptica</jats:italic>, which are respiratory mucosal pathogens of mammals, produce and utilize the siderophore alcaligin to acquire iron in response to iron starvation. A predicted permease of the major facilitator superfamily class of membrane efflux pumps, AlcS (synonyms, OrfX and Bcr), was reported to be encoded within the alcaligin gene cluster. In this study,<jats:italic>alcS</jats:italic>null mutants were found to be defective in growth under iron starvation conditions, in iron source utilization, and in alcaligin export.<jats:italic>trans</jats:italic>complementation using cloned<jats:italic>alcS</jats:italic>genes of<jats:italic>B. pertussis</jats:italic>or<jats:italic>B. bronchiseptica</jats:italic>restored the wild-type phenotype to the<jats:italic>alcS</jats:italic>mutants. Although the levels of extracellular alcaligin measured in<jats:italic>alcS</jats:italic>strain culture fluids were severely reduced compared with the wild-type levels,<jats:italic>alcS</jats:italic>mutants had elevated levels of cell-associated alcaligin, implicating AlcS in alcaligin export. Interestingly, a Δ<jats:italic>alcA</jats:italic>mutation that eliminated alcaligin production suppressed the growth defects of<jats:italic>alcS</jats:italic>mutants. This suppression and the alcaligin production defect were reversed by<jats:italic>trans</jats:italic>complementation of the Δ<jats:italic>alcA</jats:italic>mutation in the double-mutant strain, confirming that the growth-defective phenotype of<jats:italic>alcS</jats:italic>mutants is associated with alcaligin production. In an<jats:italic>alcA</jats:italic>::mini-Tn<jats:italic>5 lacZ1</jats:italic>operon fusion strain background, an<jats:italic>alcS</jats:italic>null mutation resulted in enhanced AlcR-dependent transcriptional responsiveness to alcaligin inducer; conversely, AlcS overproduction blunted the transcriptional response to alcaligin. These transcription studies indicate that the alcaligin exporter activity of AlcS is required to maintain appropriate intracellular alcaligin levels for normal inducer sensing and responsiveness necessary for positive regulation of alcaligin system gene expression.</jats:p>

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