Multiple Factors Independently Regulate \u3ci\u3ehilA\u3c/i\u3e and Invasion Gene Expression in \u3ci\u3eSalmonella enterica\u3c/i\u3e Serovar Typhimurium

HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the PhoR-PhoB two-component regulatory system, suggesting that hilA expression may be repressed by PhoRPhoB under low extracellular inorganic phosphate conditions. FadD is required for uptake and degradation of long-chain fatty acids, and our analysis of the fadD mutant indicates that hilA is regulated by a FadDdependent, FadR-independent mechanism. Thus, fatty acid derivatives may act as intracellular signals to regulate hilA expression. flhDC and fliA encode transcription factors required for flagellum production, motility, and chemotaxis. Complementation studies with flhC and fliA mutants indicate that FliZ, which is encoded in an operon with fliA, activates expression of hilA, linking regulation of hilA with motility. Finally, epistasis tests showed that PhoB, FadD, FliZ, SirA, and EnvZ act independently to regulate hilA expression and invasion. In summary, our screen has identified several distinct pathways that can modulate S. enterica serovar Typhimurium’s ability to express hilA and invade host cells. Integration of signals from these different pathways may help restrict invasion gene expression during infection

Multiple Factors Independently Regulate \u3ci\u3ehilA\u3c/i\u3e and Invasion Gene Expression in \u3ci\u3eSalmonella enterica\u3c/i\u3e Serovar Typhimurium

HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the PhoR-PhoB two-component regulatory system, suggesting that hilA expression may be repressed by PhoRPhoB under low extracellular inorganic phosphate conditions. FadD is required for uptake and degradation of long-chain fatty acids, and our analysis of the fadD mutant indicates that hilA is regulated by a FadDdependent, FadR-independent mechanism. Thus, fatty acid derivatives may act as intracellular signals to regulate hilA expression. flhDC and fliA encode transcription factors required for flagellum production, motility, and chemotaxis. Complementation studies with flhC and fliA mutants indicate that FliZ, which is encoded in an operon with fliA, activates expression of hilA, linking regulation of hilA with motility. Finally, epistasis tests showed that PhoB, FadD, FliZ, SirA, and EnvZ act independently to regulate hilA expression and invasion. In summary, our screen has identified several distinct pathways that can modulate S. enterica serovar Typhimurium’s ability to express hilA and invade host cells. Integration of signals from these different pathways may help restrict invasion gene expression during infection

Identification, Timing, and Signal Specificity of Pseudomonas aeruginosa Quorum-Controlled Genes: a Transcriptome Analysis

There are two interrelated acyl-homoserine lactone quorum-sensing-signaling systems in Pseudomonas aeruginosa. These systems, the LasR-LasI system and the RhlR-RhlI system, are global regulators of gene expression. We performed a transcriptome analysis to identify quorum-sensing-controlled genes and to better understand quorum-sensing control of P. aeruginosa gene expression. We compared gene expression in a LasI-RhlI signal mutant grown with added signals to gene expression without added signals, and we compared a LasR-RhlR signal receptor mutant to its parent. In all, we identified 315 quorum-induced and 38 quorum-repressed genes, representing about 6% of the P. aeruginosa genome. The quorum-repressed genes were activated in the stationary phase in quorum-sensing mutants but were not activated in the parent strain. The analysis of quorum-induced genes suggests that the signal specificities are on a continuum and that the timing of gene expression is on a continuum (some genes are induced early in growth, most genes are induced at the transition from the logarithmic phase to the stationary phase, and some genes are induced during the stationary phase). In general, timing was not related to signal concentration. We suggest that the level of the signal receptor, LasR, is a critical trigger for quorum-activated gene expression. Acyl-homoserine lactone quorum sensing appears to be a system that allows ordered expression of hundreds of genes during P. aeruginosa growth in culture

Multiple Factors Independently Regulate \u3ci\u3ehilA\u3c/i\u3e and Invasion Gene Expression in \u3ci\u3eSalmonella enterica\u3c/i\u3e Serovar Typhimurium

HilA activates the expression of Salmonella enterica serovar Typhimurium invasion genes. To learn more about regulation of hilA, we isolated Tn5 mutants exhibiting reduced hilA and/or invasion gene expression. In addition to expected mutations, we identified Tn5 insertions in pstS, fadD, flhD, flhC, and fliA. Analysis of the pstS mutant indicates that hilA and invasion genes are repressed by the response regulator PhoB in the absence of the Pst high-affinity inorganic phosphate uptake system. This system is required for negative control of the PhoR-PhoB two-component regulatory system, suggesting that hilA expression may be repressed by PhoRPhoB under low extracellular inorganic phosphate conditions. FadD is required for uptake and degradation of long-chain fatty acids, and our analysis of the fadD mutant indicates that hilA is regulated by a FadDdependent, FadR-independent mechanism. Thus, fatty acid derivatives may act as intracellular signals to regulate hilA expression. flhDC and fliA encode transcription factors required for flagellum production, motility, and chemotaxis. Complementation studies with flhC and fliA mutants indicate that FliZ, which is encoded in an operon with fliA, activates expression of hilA, linking regulation of hilA with motility. Finally, epistasis tests showed that PhoB, FadD, FliZ, SirA, and EnvZ act independently to regulate hilA expression and invasion. In summary, our screen has identified several distinct pathways that can modulate S. enterica serovar Typhimurium’s ability to express hilA and invade host cells. Integration of signals from these different pathways may help restrict invasion gene expression during infection

Gene-Swapping Mediates Host Specificity among Symbiotic Bacteria in a Beneficial Symbiosis

<div><p>Environmentally acquired beneficial associations are comprised of a wide variety of symbiotic species that vary both genetically and phenotypically, and therefore have differential colonization abilities, even when symbionts are of the same species. Strain variation is common among conspecific hosts, where subtle differences can lead to competitive exclusion between closely related strains. One example where symbiont specificity is observed is in the sepiolid squid-<i>Vibrio</i> mutualism, where competitive dominance exists among <i>V. fischeri</i> isolates due to subtle genetic differences between strains. Although key symbiotic loci are responsible for the establishment of this association, the genetic mechanisms that dictate strain specificity are not fully understood. We examined several symbiotic loci (<i>lux</i>-bioluminescence, <i>pil</i> = pili, and <i>msh</i>-mannose sensitive hemagglutinin) from mutualistic <i>V. fischeri</i> strains isolated from two geographically distinct squid host species (<i>Euprymna tasmanica</i>-Australia and <i>E. scolopes</i>-Hawaii) to determine whether slight genetic differences regulated host specificity. Through colonization studies performed in naïve squid hatchlings from both hosts, we found that all loci examined are important for specificity and host recognition. Complementation of null mutations in non-native <i>V. fischeri</i> with loci from the native <i>V. fischeri</i> caused a gain in fitness, resulting in competitive dominance in the non-native host. The competitive ability of these symbiotic loci depended upon the locus tested and the specific squid species in which colonization was measured. Our results demonstrate that multiple bacterial genetic elements can determine <i>V. fischeri</i> strain specificity between two closely related squid hosts, indicating how important genetic variation is for regulating conspecific beneficial interactions that are acquired from the environment.</p></div

<i>lux</i> operon data.

<p>Colonization assays 48-hour post-infection of juvenile (A) <i>Euprymna scolopes</i> and (B) <i>Euprymna tasmanica</i> by their respective wild-type (ES114 or ETJB1H), mutant, and complement strains of the <i>lux</i> operon for <i>Vibrio fischeri</i>. Infection efficiency data is plotted as the log values of the relative competitiveness index (RCIs), calculated by dividing the ratio of mutant to wild-type by the starting ratio <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0101691#pone.0101691-Hussa1" target="_blank">[28]</a>. If the RCI is <1 the mutant strain was outcompeted by the wild-type, the wild-type strain was outcompeted by the mutant if the value is >1, and a RCI equal to 1 indicates no competitive difference. Data points represent individual animals and the position of the figures on the y axis is merely for spacing. Vertical line represents the median value of each data plot.</p

<i>msh</i> operon data.

<p>Colonization assays 48-hour post-infection of juvenile (A) <i>Euprymna scolopes</i> and (B) <i>Euprymna tasmanica</i> by their respective wild-type (ES114 or ETJB1H), mutant, and complement strains of <i>msh</i> genes for <i>Vibrio fischeri</i>. Infection efficiency data is plotted as the log values of the relative competitiveness index (RCIs), calculated by dividing the ratio of mutant to wild-type by the starting ratio <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0101691#pone.0101691-Hussa1" target="_blank">[28]</a>. If the RCI is <1 the mutant strain was outcompeted by the wild-type, the wild-type strain was outcompeted by the mutant if the value is >1, and a RCI equal to 1 indicates no competitive difference. Data points represent individual animals and the position of the figures on the y axis is merely for spacing. Vertical line represents the median value of each data plot.</p