Central metabolism controls transcription of a virulence gene regulator in Vibrio cholerae.

Oregon State University
Microbiology (Impact Factor: 2.84). 02/2013; DOI: 10.1099/mic.0.064865-0
Source: PubMed

ABSTRACT ToxT is the central regulatory protein involved in activation of the main virulence genes in Vibrio cholerae. We have identified transposon insertions in central metabolism genes, whose disruption increases toxT transcription. These disrupted genes encode the primary respiration-linked sodium pump (NADH-ubiquinone oxidoreductase or NQR) and certain tricarboxylic acid (TCA) cycle enzymes. Observations made following stimulation of respiration in the nqr mutant or chemical inhibition of NQR activity in the TCA cycle mutants led to the hypothesis that NQR affects toxT transcription via the TCA cycle. That toxT transcription increased when the growth medium was supplemented with citrate, but decreased with oxaloacetate, focused our attention on the TCA cycle substrate acetyl-coenzyme A (acetyl-CoA) and its non-TCA cycle metabolism. Indeed, both the nqr and TCA cycle mutants increased acetate excretion. A similar correlation between acetate excretion and toxT transcription was observed in the tolC mutant and upon amino acid (NRES) supplementation. Since acetate and its tendency to decrease pH exerted no strong effect on toxT transcription, and since disruption of the major acetate excretion pathway increased toxT transcription, we propose that toxT transcription is regulated by either acetyl-CoA or some close derivative.

1 Follower
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: The Na+ translocating NADH:quinone oxidoreductase (Na+-NQR) is a unique respiratory enzyme catalyzing the electron transfer from NADH to quinone coupled with the translocation of sodium ions across the membrane. Typically, Vibrio spp., including Vibrio cholerae, have this enzyme but lack the proton-pumping NADH:ubiquinone oxidoreductase (Complex I). Thus, Na+-NQR should significantly contribute to multiple aspects of V. cholerae physiology; however, no detailed characterization of this aspect has been reported so far. In this study, we broadly investigated the effects of loss of Na+-NQR on V. cholerae physiology by using Phenotype Microarray (Biolog), transcriptome and metabolomics analyses. We found that the V. cholerae ΔnqrA-F mutant showed multiple defects in metabolism detected by Phenotype Microarray. Transcriptome analysis revealed that the V. cholerae ΔnqrA-F mutant up-regulates 31 genes and down-regulates 55 genes in both early and mid-growth phases. The most up-regulated genes included the cadA and cadB genes, encoding a lysine decarboxylase and a lysine/cadaverine antiporter, respectively. Increased CadAB activity was further suggested by the metabolomics analysis. The down-regulated genes include sialic acid catabolism genes. Metabolomic analysis also suggested increased reductive pathway of TCA cycle and decreased purine metabolism in the V. cholerae ΔnqrA-F mutant. Lack of Na+-NQR did not affect any of the Na+ pumping-related phenotypes of V. cholerae suggesting that other secondary Na+ pump(s) can compensate for Na+ pumping activity of Na+-NQR. Overall, our study provides important insights into the contribution of Na+-NQR to V. cholerae physiology.
    PLoS ONE 05/2014; 9(5):e97083. DOI:10.1371/journal.pone.0097083 · 3.53 Impact Factor
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Classical studies have focused on the role that individual regulators play in controlling virulence gene expression. An emerging theme, however, is that bacterial metabolism also plays a key role in this process. Our previous work identified a series of proteins that were implicated in the regulation of virulence. One of these proteins was AdhE, a bi-functional acetaldehyde-CoA dehydrogenase and alcohol dehydrogenase. Deletion of its gene (adhE) resulted in elevated levels of extracellular acetate and a stark pleiotropic phenotype: strong suppression of the Type Three Secretion System (T3SS) and overexpression of non-functional flagella. Correspondingly, the adhE mutant bound poorly to host cells and was unable to swim. Furthermore, the mutant was significantly less virulent than its parent when tested in vivo, which supports the hypothesis that attachment and motility are central to the colonization process. The molecular basis by which AdhE affects virulence gene regulation was found to be multifactorial, involving acetate-stimulated transcription of flagella expression and post-transcriptional regulation of the T3SS through Hfq. Our study reveals fascinating insights into the links between bacterial physiology, the expression of virulence genes, and the underlying molecular mechanism mechanisms by which these processes are regulated.
    Molecular Microbiology 05/2014; 93(1). DOI:10.1111/mmi.12651 · 5.03 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Vibrio fischeri isolated from Euprymna scolopes (Cephalopoda: Sepiolidae) was used to create 24 lines that were serially passaged through the non-native host Euprymna tasmanica for 500 generations. These derived lines were characterized for biofilm formation, swarming motility, carbon source utilization, and in vitro bioluminescence. Phenotypic assays were compared between "ES" (E. scolopes) and "ET" (E. tasmanica) V. fischeri wild isolates to determine if convergent evolution was apparent between E. tasmanica evolved lines and ET V. fischeri. Ecological diversification was observed in utilization of most carbon sources examined. Convergent evolution was evident in motility, biofilm formation, and select carbon sources displaying hyperpolymorphic usage in V. fischeri. Convergence in bioluminescence (a 2.5-fold increase in brightness) was collectively evident in the derived lines relative to the ancestor. However, dramatic changes in other properties-time points and cell densities of first light emission and maximal light output and emergence of a lag phase in growth curves of derived lines-suggest that increased light intensity per se was not the only important factor. Convergent evolution implies that gnotobiotic squid light organs subject colonizing V. fischeri to similar selection pressures. Adaptation to novel hosts appears to involve flexible microbial metabolism, establishment of biofilm and swarmer V. fischeri ecotypes, and complex changes in bioluminescence. Our data demonstrate that numerous alternate fitness optima or peaks are available to V. fischeri in host adaptive landscapes, where novel host squids serve as habitat islands. Thus, V. fischeri founder flushes occur during the initiation of light organ colonization that ultimately trigger founder effect diversification.
    Microbial Ecology 01/2014; 67(3). DOI:10.1007/s00248-013-0356-3 · 3.12 Impact Factor