ABSTRACT

Pseudomonas aeruginosa uses quorum sensing (QS) to regulate the production of a battery of secreted products. At least some of these products are shared among the population and serve as public goods. When P. aeruginosa is grown on casein as the sole carbon and energy source, the QS-induced extracellular protease elastase is required for growth. We isolated a P. aeruginosa variant, which showed increased production of QS-induced factors after repeated transfers in casein broth. This variant, P. aeruginosa QS*, had a mutation in the glutathione synthesis gene gshA. We describe several experiments that show a gshA coding variant and glutathione affect the QS response. The P. aeruginosa QS transcription factor LasR has a redox-sensitive cysteine (C79). We report that GshA variant cells with a LasR C79S substitution show a similar QS response to that of wild-type P. aeruginosa. Surprisingly, it is not LasR but the QS transcription factor RhlR that is more active in bacteria containing the variant gshA. Our results demonstrate that QS integrates information about cell density and the cellular redox state via glutathione levels.
IMPORTANCE Pseudomonas aeruginosa and other bacteria coordinate group behaviors using a chemical communication system called quorum sensing (QS). The QS system of P. aeruginosa is complex, with several regulators and signals. We show that decreased levels of glutathione lead to increased gene activation in P. aeruginosa, which did not occur in a strain carrying the redox-insensitive variant of a transcription factor. The ability of P. aeruginosa QS transcription factors to integrate information about cell density and cellular redox state shows these transcription factors can fine-tune levels of the gene products they control in response to at least two types of signals or cues.

INTRODUCTION

Pseudomonas aeruginosa uses a cell-cell communication system called quorum sensing (QS) to regulate the production of a large number of secreted and excreted factors, several of which are virulence factors. The QS system of P. aeruginosa is complex and involves several signals and receptors that ultimately regulate the transcription of hundreds of genes. Two of the QS receptors, LasR and RhlR, recognize acyl-homoserine lactone (AHL) signals and are homologs of the Vibrio fischeri receptor LuxR. These signals, N-3-oxo-dodecanoyl homoserine lactone (3OC12-HSL) and N-butyryl-homoserine lactone (C4-HSL), are synthesized by the LasI and RhlI signal synthases, respectively. LasR recognizes 3OC12-HSL and RhlR recognizes C4-HSL. P. aeruginosa also has a quinolone signal receptor, PqsR. LasR has previously been described as a “master” QS regulator, as activation of the RhlR and PqsR systems depends on LasR in laboratory strains (reviewed in references 1 to 3).
Many QS-regulated products are shared among the population, which is a type of cooperative behavior (4, 5). One such product is the protease elastase, encoded by lasB. lasB transcription is induced by both LasR and RhlR (6). When P. aeruginosa strain PAO1 is grown on minimal medium containing casein as the sole carbon and energy source, the production of elastase is required to break down casein into amino acids and peptides, which P. aeruginosa can take up and use for carbon and energy. This is a cooperative behavior, and growth on casein is susceptible to cheating. Over about 100 generations, LasR-null “social cheaters” emerge in a cooperating population (7). These cheaters produce neither elastase nor other quorum-regulated products, but they are able to avail themselves of the casein breakdown products from elastase secreted by cooperating bacteria.
The emergence of these social cheaters in cooperating P. aeruginosa populations is subject to several restrictions. For example, cooperators can produce cyanide, with the effect of preferentially impairing the growth of cheats (8, 9). Cooperators may also delay making metabolically expensive exoproducts until stationary phase (“metabolic prudence”) (10). However, there are also conditions in which the frequency of cheaters is unrestrained (11, 12).
When P. aeruginosa strain PAO1 is grown in a minimal medium where casein is the sole carbon and nitrogen source, cheaters are unrestrained, resulting in a tragedy of the commons, for lack of sufficient cooperators (11). Here, the cooperators in the population appear to have an increasing burden to generate QS-induced products. There are large increases in production of both elastase and the phenazine pyocyanin in populations repeatedly transferred on casein broth without ammonium sulfate (11). As such, it appears that there is selection for cheaters while the cooperators endure an increased cost of QS.
One potential environmental input into P. aeruginosa QS is the redox state of the cell. It has been reported that redox sensing can influence the QS regulator LasR through a cysteine at residue 79 adjacent to the ligand-binding site (13, 14). The redox state of some bacteria, including P. aeruginosa, is regulated in part by the production of glutathione, a tripeptide thiol antioxidant. In P. aeruginosa, glutathione is produced in a two-step process. gshA encodes a protein that ligates the γ-carboxyl group of glutamate to cysteine, generating γ-glutamylcysteine, which is condensed with glycine by the gshB gene product, yielding glutathione (15). The contribution of these genes to P. aeruginosa pathogenesis appears to depend on the setting. Deletion of either gshA or gshB has been shown to reduce the virulence of this bacterium in a Caenorhabditis elegans model (16); however, in a murine wound model, gshA (but not gshB) transposon mutants were enriched, suggesting that gshA deletions might be more virulent than the wild type (WT) (17).
We have isolated a stable P. aeruginosa variant, QS*, which overproduces protease and pyocyanin from a population repeatedly transferred in casein broth where casein is the sole carbon and nitrogen source. We discovered that the genome of QS* had a single-nucleotide substitution in gshA. This gshA mutation contributes to enhanced production of QS-regulated products. We present evidence that QS integrates information about cell population density with information about glutathione levels or cellular redox states.

RESULTS

Isolation of a stable QS-hyperactive mutant.

We previously reported (11) that when P. aeruginosa PAO1 is subcultured through about 50 doublings in casein broth without added ammonium sulfate, production of pyocyanin and elastase increases dramatically compared to that in the WT grown in casein broth with added ammonium sulfate. We did not isolate evolved strains in the previous study (11). Here, we asked if variants with highly active QS systems can be isolated from populations after 15 days of passage in casein broth where casein is the sole carbon and nitrogen source. When isolates were spotted on milk agar plates, they showed some variation in terms of protease production as assessed by the size of the zones of clearance around the colony (Fig. 1A and B). We chose one isolate that produced particularly large zones of clearance for further study. This isolate, which we call P. aeruginosa QS*, also produced high levels of the QS-induced pigment pyocyanin when grown in casein broth (Fig. 1C).
FIG 1
FIG 1 The QS-hyperactive phenotype. (A) Isolates from day 15 passage in casein broth without ammonium sulfate plated on skim milk agar for 24 h. The top row, from left to right, includes wild-type P. aeruginosa PAO1, QS*, and a P. aeruginosa PAO1 lasR deletion mutant. The bottom two rows include eight different isolates (each of which has an inactivating psdR mutation) from the day 15 passage. (B) The relative size of zones of clearance in panel A compared to PAO1 (100%). (C) Cultures of P. aeruginosa PAO1, the evolved QS* mutant, a PsdR-null variant, and a GshAT44P-PsdR-null double variant grown in casein broth for 16 h. The QS* culture is visibly blue as a result of pyocyanin overproduction. The GshAT44P-PsdR-null double mutant has a phenotype similar to that of QS*.
We performed whole-genome sequencing of QS* in an effort to identify the genetic basis of its highly active QS system. Isolates adapted to growth in casein broth, such as those shown in Fig. 1A, carry mutations in psdR, which codes for a repressor of amino acid uptake genes (18). An inactivating mutation in psdR results in improved growth in casein broth (18). Not surprisingly, QS* had a psdR promoter mutation. We also found an A130C mutation in gshA. This A130C mutation codes for a glutamine-cysteine ligase with a T44P substitution in the GshA protein sequence. The glutamine-cysteine ligase catalyzes an essential step in glutathione synthesis (19). However, it was unclear to us how the combined two mutations contribute to the QS-hyperactive phenotype.

A gshA mutation contributes to the QS* phenotype.

To test whether the gshA A130C substitution (which encodes the T44P amino acid change, GshAT44P), in combination with the psdR mutation, is responsible for the QS* phenotype, we engineered a PsdR-null mutation in a gshA A130C substitution background and compared it to a PsdR-null strain. The GshAT44P-PsdR-null double variant appeared to be similar to QS* after overnight growth in casein broth (Fig. 1C). The PsdR-null variant alone, however, was less blue than either QS* or the GshAT44P-PsdR-null double variant. These results suggest that gshA mutation and psdR mutation together confer the QS* phenotype. Since the function of PsdR in casein broth has been resolved (18), we were curious about how the GshAT44P variant contributes to the QS* phenotype.
We asked if either the GshAT44P variant or a GshA-null variant affected growth compared to that of the WT. Growth of a strain containing the GshAT44P variant in Casamino Acids (CAA) broth was indistinguishable from growth of the WT and the QS* mutant (Fig. 2A). CAA broth simulates the nutritional condition of casein broth without the requirement for QS-regulated proteases to enable growth. However, a mutant with a deletion of gshA (GshA-null) exhibited slower growth than that of other strains (Fig. 2A). The impaired growth of the deletion mutant suggested that the GshAT44P variant retains some functionality. In contrast to growth in CAA broth, there were clear differences between the strains in casein broth. The double variant and QS* exhibited substantially higher growth rates than either the GshAT44P variant or the WT (Fig. 2B).
FIG 2
FIG 2 Growth of Pseudomonas aeruginosa PAO1 and various mutants within 24 h in CAA broth (A) and casein broth (B). Growth in CAA broth was measured as optical density at 600 nm. Growth in casein broth was measured as CFU per milliliter because, as casein is digested, intermediates interfere with optical density measurement. Wild-type P. aeruginosa PAO1, the evolved isolate QS*, the GshA-null mutant, the engineered GshAT44P mutant, and the engineered GshAT44P-PsdR mutant are shown. Data are shown as the mean ± range from two independent experiments.
We measured production of pyocyanin, elastase, and the QS signals 3OC12-HSL and C4-HSL by QS*, GshAT44P, and the WT PAO1 in casein broth without ammonium sulfate (Fig. 3). QS* produced more pyocyanin than the wild type and GshAT44P (Fig. 3A) throughout growth, and protease and signal production were earlier in QS* than in other strains (Fig. 3B to D). This phenotype of QS* is likely attributable in part to its faster growth in casein broth (Fig. 2B). The GshAT44P variant also produced more pyocyanin and C4-HSL in growth than the WT (Fig. 3A and D). However, protease and 3OC12-HSL production were similar between the WT and the GshAT44P variant (Fig. 3B and C). Together, these data support the view that the gshA mutation and the psdR mutation in tandem are responsible for the QS* phenotype.
FIG 3
FIG 3 Early and elevated production of QS-activated products in P. aeruginosa QS* and other GshA mutants. Pyocyanin (A), protease (B), 3OC12-HSL (C), and C4-HSL (D) levels from the wild type (PAO1), QS*, and the GshAT44P mutant are shown. Bacteria were grown in casein broth for 0, 6, 12, 18, and 24 h. Data are shown as the mean ± standard error of the mean (SEM) from four independent experiments. *, P < 0.05; **, P < 0.01 in comparisons between QS* and PAO1. #, P < 0.05; ##, P < 0.01 for comparisons between GshAT44P and PAO1 by two-way analysis of variance (ANOVA).
We then asked whether adding glutathione or cysteine to casein broth might phenotypically complement both QS* and the GshAT44P variant. In fact, glutathione addition significantly reduced pyocyanin production (Fig. 4) by both QS* and strain GshAT44P. Cysteine addition also reduced the pyocyanin production of GshAT44P (see Fig. S1 in the supplemental material). These experiments are consistent with the conclusion that gshA mutations result in hyperactive QS responses, representing a cellular response to glutathione.
FIG 4
FIG 4 Growth of P. aeruginosa wild type and GshA mutants in casein broth with added glutathione. (A) The wild type (PAO1), QS*, and the engineered GshAT44P mutant were grown in casein broth with (+) or without (−) 200 µM glutathione for 18 h. (B) Pyocyanin production for PAO1, QS*, and GshAT44P in casein broth with blank or with added 200 µM glutathione for 18 h. Data are shown as the mean ± SEM from three independent experiments. *, P < 0.05 by the Student’s t test.

The GshAT44P variant has lower levels of glutathione.

As discussed above, we suspected that the GshAT44P enzyme is not completely inactive. To address this possibility, we measured total and reduced glutathione levels in WT PAO1, the QS* isolate, the engineered GshAT44P mutant, and the gshA deletion mutant. Both the QS* and GshAT44P variants had lower levels of total glutathione than the WT and higher levels of total glutathione than the GshA-null mutant (Fig. 5A). There was also a statistically significant difference in reduced glutathione levels between the WT and the GshA-null mutant (Fig. 5B). Furthermore, the decrease in glutathione levels was reversed by expression of the WT gshA allele on a plasmid (Fig. 5C and D).
FIG 5
FIG 5 The effect of gshA mutations on cellular glutathione levels. (A and B) Total (A) and reduced (B) glutathione levels in the WT PAO1, QS*, the GshAT44P mutant, and the GshA-null mutant. (C and D) Total (C) and reduced (D) glutathione levels in the WT, the GshA deletion mutant containing pBBR1MCS5, and the GshA deletion mutant containing pBBR1MCS5-gshA. Cells were grown in Casamino Acids broth and sampled at an OD600 of 0.5. Glutathione was measured as described in Materials and Methods. Data are shown as the mean ± SEM from three independent experiments. *, P < 0.05; **, P < 0.01 by one-way ANOVA.

QS* containing a redox-insensitive variant of LasR does not have the gshA mutation phenotype.

A previous report (13) identified a cysteine at residue 79 in LasR as redox sensitive. This cysteine is adjacent to the signal-binding pocket of LasR (20), and its replacement by serine yields a functional LasR that is not affected by oxidation (13, 14). We reasoned that either QS* or the GshAT44P variant containing the LasRC79S variant would not exhibit a hyperactive QS phenotype. The QS*-LasRC79S strain produced significantly less pyocyanin than QS*, and pyocyanin production by GshAT44P-LasRC79S was similar to that of the WT (Fig. 6). Therefore, abrogating the redox sensitivity of LasR reverses the QS* phenotype.
FIG 6
FIG 6 The QS* response to glutathione requires LasR C79. (A) Images of 18-h casein broth cultures. Protease production has resulted in a chalky appearance of the broth due to partial casein digestion. The wild-type PAO1, QS*, and the engineered GshA T44P mutant (GshAT44P) in a LasR WT background or in a LasR C79S mutant background (LasRC79S) are shown. (B) Pyocyanin production of the same strains after 18 h of growth. Data are shown as the mean ± SEM from three independent experiments. *, P < 0.05; **, P < 0.01 by one-way ANOVA.

The gshA variant and RhlR-activated products.

We hypothesized that decreased glutathione in QS* might result in higher levels of LasR-regulated gene expression. To test this hypothesis, we monitored transcription from the lasI promoter in wild-type PAO1, QS*, and the engineered GshAT44P mutant by using an lasI-gfp fusion (Fig. 7A). Surprisingly, even though total glutathione levels are diminished in QS* and GshAT44P, there was no difference in green fluorescent protein (GFP) accumulation among these three strains. This result paradoxically suggested that redox sensing by LasR, as shown in Fig. 6, may not be the primary mechanism by which the glutathione effect is mediated and that the phenotype of the dual GshAT44P-LasRC79S variant might reflect a dominant effect of the LasR variant.
FIG 7
FIG 7 The influence of the GshAT44P mutant polypeptide on transcription from an LasR-responsive and RhlR-responsive promoter. GFP fluorescence was measured from wild-type PAO1, the QS* variant, the GshAT44P mutant, and the LasR-null RhlR-null double mutant containing either a lasI-gfp reporter (A) or a rhlA-gfp reporter (B). Cells were grown in CAA medium. Data are shown as the mean ± SEM from four independent experiments. *, P < 0.05; **, P < 0.01 in comparisons between QS* and PAO1; ##, P < 0.01 for comparisons between GshAT44P and PAO1 by two-way ANOVA.
We reasoned that the impact of the GshAT44P variant might be to change the contributions of LasR and RhlR to QS gene activation, favoring RhlR, which is not a redox-active polypeptide (13). We measured transcription of gfp driven from the RhlR-dependent rhlA promoter. Unlike transcription from the lasI promoter, transcription from this RhlR-dependent promoter is significantly elevated in QS* and GshAT44P compared to that of the WT (Fig. 7B). This result suggests that there may be additional mechanisms, other than through the C79 residue of LasR, by which glutathione modulates QS gene activation. It is, however, consistent with the observation of early C4-HSL production by QS* and the GshAT44P mutant (Fig. 3D). Somehow, a 50% reduction of total glutathione (Fig. 5A) allows for increased RhlR activity, possibly by accelerated production of C4-HSL (Fig. 3D). There may be an impact on early activation of LasR that is not resolved in our experiments, or it may be that there is a second mechanism involving RhlR-I expression or both. The ultimate effect is that in QS*, there is earlier production of protease (which is induced primarily by LasR but also by RhlR) and greater production of pyocyanin (RhlR regulated) than in PAO1.
Could redox sensitivity of QS signal receptors be common in AHL QS systems? We aligned LasR to other LuxR homologs and found that a cysteine residue is conserved in several but not all of these transcription factors (see Fig. S2 in the supplemental material). The other P. aeruginosa QS transcription factors, RhlR and QscR, do not have a cysteine residue aligning with LasR C79, which is consistent with the prior report that neither was reactive in a global study of redox-reactive P. aeruginosa polypeptides (13).

DISCUSSION

We showed previously that during serial passage of P. aeruginosa PAO1 in minimal medium with casein as the sole carbon, energy, and nitrogen source, the populations evolve to produce increased amounts of the QS-induced pyocyanin and extracellular proteases (11). To further our understanding of this phenomenon, we isolated individuals from a culture after 15 daily transfers in casein broth without added ammonium sulfate. There was diversity in terms of clonal production of proteases on skim milk agar plates. We chose one particularly strong protease producer for further study and discovered that a single nucleotide mutation in gshA could partially account for the enhanced protease production. This mutation also results in enhanced production of pyocyanin and C4-HSL at 18 h and 24 h compared to that of the WT. We have called this variant QS*. Both QS* and an engineered P. aeruginosa strain with the QS* nucleotide change in gshA show reduced cellular levels of glutathione, and addition of glutathione to casein broth markedly inhibits pyocyanin production. Our results reinforce the idea that there is a connection between QS and redox sensing in P. aeruginosa (13).
A recent report showed that a GshA-null mutant produces less pyocyanin and biofilm than its parent (21). There was no obvious difference in QS signal production (21). These results, along with ours demonstrating a growth defect in the gshA deletion strain (Fig. 2), suggest that the QS* phenotype is partially a consequence of the specific amino acid substitution (T44P) in GshA and does not result from complete loss of GshA activity. We propose that the hyperactive QS phenotype occurs because the decrease in glutathione levels from the T44P variant of GshA modulates QS, with the ultimate outcome that RhlR-regulated gene activation is increased compared to that in PAO1 (Fig. 7), although the mechanism by which this happens is unknown.
Deng et al. (13) showed that LasR is a redox-active protein. The cysteine at residue 79 of LasR was identified as redox sensitive, and addition of H2O2 to growing P. aeruginosa cells resulted in lower LasR activity, as measured by an lasI-lacZ fusion reporter. Because C79 is adjacent to the AHL-binding pocket, it may influence the binding of LasR to its signal 3OC12-HSL (20). In our experiments, the gshA variant conferred a hyperactive QS phenotype, and this was abrogated in a strain with an LasR C79S substitution (Fig. 6). These data suggest that there may be an interaction between glutathione and LasR (Fig. 6), and the results show the importance of glutathione in the level of QS response.
The connection between QS and redox sensing adds to our knowledge of the interplay between QS and other aspects of the physiology of P. aeruginosa. This extensive regulation of QS in this species probably reflects a high potential for adapting to diverse environments (22). Our finding that a coding variant of gshA results in increased production of various secreted products expands the model of QS regulation in P. aeruginosa. Multiple environmental inputs can modulate QS in this species, including redox state via cellular glutathione levels.

MATERIALS AND METHODS

Bacterial strains and culture conditions.

Strains and plasmids used in this study are listed in Table S1 in the supplemental material. All P. aeruginosa strains were derived from the wild-type PAO1-UW (22). Bacteria were grown in LB broth buffered with 50 mM 3-(N-morpholino)propanesulfonic acid (LB-MOPS), pH 7.0, or in photosynthesis medium (23) without ammonium sulfate supplemented with 1% (wt/vol) sodium caseinate (casein broth) or 1% (wt/vol) Casamino Acids (CAA broth). Bacteria were grown at 37°C with shaking (250 rpm) in 18-mm glass tubes containing 4 or 5 ml of broth. Agar plates contained 1.5% agar.
Experiments in casein broth were started from 18-h LB-MOPS broth cultures that had been inoculated with a freshly grown single colony of P. aeruginosa. The initial optical density at 600 nm (OD600) was 0.025, unless otherwise specificed.

Construction of mutant strains.

We used a homologous recombination approach (24, 25) to generate mutations in lasR, gshA, or psdR. Briefly, sequence-verified PCR products containing a variant or knockout allele and PCR-amplified pEXG2 vector (26) were used to transform Escherichia coli DH5α. Transformants were isolated by selective plating and confirmed by PCR. We miniprepped the constructed plasmids and used the plasmid preparations to electrotransform E. coli S17. The E. coli isolates were then mated with P. aeruginosa isolates (24). We selected transconjugants by plating on Pseudomonas isolation agar containing gentamicin (100 μg/ml) and counterselected on no-salt LB agar containing 10% sucrose (24). All mutations were confirmed by Sanger sequencing of the allele. Primers used for amplification and construction of plasmids are listed in Table S2 in the supplemental material. Gentamicin (10 to 30 µg/ml for E. coli; 100 μg/ml for P. aeruginosa) was used for plasmid maintenance.

Whole-genome sequencing.

Genomic DNA was isolated from bacteria grown in LB-MOPS broth by using Puregene yeast/bacteria kit B (Qiagen). DNA libraries were prepared by using a Nextera kit (Illumina), and DNA was sequenced with an Illumina MiSeq. Variant analysis was performed as previously described by using Strand NGS version 2.9 software (Strand Life Sciences, Bangalore, India) (27).

Glutathione measurements.

Bacteria grown overnight in LB-MOPS broth were used to inoculate CAA broth (initial OD600 of 0.05 or 0.1). Glutathione was measured as described elsewhere (28). Briefly, cells were pelleted by centrifugation and suspended in phosphate-buffered saline (PBS). The cell suspension was mixed 1:1 with 10% 5-sulfosalicylic acid, incubated on ice for 10 min, and then sonicated. For measurement of total glutathione, 40 μl of Tris(2-carboxyethyl)phosphine hydrochloride (TCEP) was added or 40 μl H2O for reduced glutathione. Samples were centrifuged at 15,600 × g for 2 min, and 25 μl of the supernatant fluid was added to 100 μl of a solution of 0.2 M N-ethylmorpholine and 0.02 M NaOH. Following the addition of 50 μl 0.5 N NaOH, glutathione was derivatized by the addition of 10 μl of 10 mM naphthalene-2,3-dicarboxaldehyde (NDA). The mixture was incubated at room temperature for 30 min, and the fluorescence intensity of the NDA-glutathione conjugate was measured at excitation wavelength (λex) 472 and emission wavelength (λem) 528. A standard curve was constructed with NDA-conjugated glutathione.

Protease and pyocyanin measurements.

We inoculated casein broth with cells from an overnight LB-MOPS broth culture and measured protease and pyocyanin at the indicated times after inoculation. Protease activity was measured with a fluorescent protease activity kit (Thermo Fisher Scientific) (18). Pyocyanin was extracted with chloroform and 0.2 N HCl as previously described (29), and the amount of pyocyanin was determined by measuring the absorbance of the aqueous phase at 520 nm.

AHL measurements.

AHLs were extracted with ethyl acetate as previously described (30). We used bioassays to measure 3OC12-HSL and C4-HSL in the ethyl acetate extracts. As described elsewhere, E. coli DH5α containing pJN105L and pSC11 was used to measure 3OC12-HSL (31). For the C4-HSL bioassays, we used E. coli containing pOHC4, an rhlA-mCherry reporter, as previously described (32).

LasR and RhlR activity measurements.

We electrotransformed strains with a plasmid containing an lasI-gfp reporter (pBS351) or an rhlA-gfp reporter (pJF01), as previously described (33). These constructs are transcriptional fusions; transcription of gfp is driven from the lasI or rhlA promoter, but gfp has its own ribosomal binding and translation start sites. Bacteria were grown to logarithmic phase in CAA broth, (OD600 = 0.2 to 0.4) and back-diluted to an OD600 of 0.005 (also in CAA broth). GFP fluorescence and OD were measured at 2-h intervals using a microplate reader.

Data availability.

Sequence data for QS* (SAMN09666232) and PAO1 (SAMN09671539) are deposited at the NCBI Sequence Read Archive.

ACKNOWLEDGMENTS

We thank Colin C. White for assistance with the glutathione measurements and John Feltner for helpful discussions.
This research was supported by National Institutes of Health grants GM059026 (to E.P.G.) and GM125714 (to A.A.D.). A.A.D. is also supported by a Career Award for Medical Scientists from the Burroughs-Wellcome Fund. Hui Zhou was supported by China Scholarship Council Grant 201606320198 for International Exchange of Personnel. Meizhen Wang was supported by China Scholarship Council Grant 201308330023 and Zhejiang Association for International Exchange of Personnel. Maxim Kostylev was supported by a fellowship from the Cystic Fibrosis Foundation (KOSTYL17F0).

Supplemental Material

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Information & Contributors

Information

Published In

cover image Journal of Bacteriology
Journal of Bacteriology
Volume 201Number 91 May 2019
eLocator: 10.1128/jb.00685-18
Editor: George O'Toole, Geisel School of Medicine at Dartmouth

History

Received: 5 November 2018
Accepted: 7 February 2019
Published online: 9 April 2019

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Keywords

  1. LasR
  2. RhlR
  3. cooperation
  4. redox sensing
  5. sociomicrobiology

Contributors

Authors

Hui Zhou
Department of Infectious Diseases, The Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China
Department of Microbiology, University of Washington, Seattle, Washington, USA
Meizhen Wang
School of Environmental Science and Engineering, Zhejiang Gongshang University, Hangzhou, China
Nicole E. Smalley
Department of Medicine, University of Washington, Seattle, Washington, USA
Maxim Kostylev
Department of Microbiology, University of Washington, Seattle, Washington, USA
Amy L. Schaefer
Department of Microbiology, University of Washington, Seattle, Washington, USA
Department of Microbiology, University of Washington, Seattle, Washington, USA
Department of Medicine, University of Washington, Seattle, Washington, USA
Department of Microbiology, University of Washington, Seattle, Washington, USA
Feng Xu
Department of Infectious Diseases, The Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, China

Editor

George O'Toole
Editor
Geisel School of Medicine at Dartmouth

Notes

Address correspondence to Ajai A. Dandekar, [email protected], or Feng Xu, [email protected].

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