N-dodecanoyl-homoserine lactone influences the levels of thiol and proteins related to oxidation-reduction process in Salmonella

Abstract

Quorum sensing is a cell-cell communication mechanism mediated by chemical signals that leads to differential gene expression in response to high population density. Salmonella is unable to synthesize the autoinducer-1 (AI-1), N-acyl homoserine lactone (AHL), but is able to recognize AHLs produced by other microorganisms through SdiA protein. This study aimed to evaluate the fatty acid and protein profiles of Salmonella enterica serovar Enteritidis PT4 578 throughout time of cultivation in the presence of AHL. The presence of N-dodecanoyl-homoserine lactone (C12-HSL) altered the fatty acid and protein profiles of Salmonella cultivated during 4, 6, 7, 12 and 36 h in anaerobic condition. The profiles of Salmonella Enteritidis at logarithmic phase of growth (4 h of cultivation), in the presence of C12-HSL, were similar to those of cells at late stationary phase (36 h). In addition, there was less variation in both protein and fatty acid profiles along growth, suggesting that this quorum sensing signal anticipated a stationary phase response. The presence of C12-HSL increased the abundance of thiol related proteins such as Tpx, Q7CR42, Q8ZP25, YfgD, AhpC, NfsB, YdhD and TrxA, as well as the levels of free cellular thiol after 6 h of cultivation, suggesting that these cells have greater potential to resist oxidative stress. Additionally, the LuxS protein which synthesizes the AI-2 signaling molecule was differentially abundant in the presence of C12-HSL. The NfsB protein had its abundance increased in the presence of C12-HSL at all evaluated times, which is a suggestion that the cells may be susceptible to the action of nitrofurans or that AHLs present some toxicity. Overall, the presence of C12-HSL altered important pathways related to oxidative stress and stationary phase response in Salmonella.

Fig 1. PCA analysis of fatty acids from Salmonella Enteritidis PT4 578 anaerobically cultivated in TSB at 37 °C in the presence or absence of C12-HSL.

PCA of the percentages of each triplicate of fatty acid profile of Salmonella in the absence (control) and presence of C12-HSL.

Fig 2. PCA, heatmap and dendrogram analyses of proteins from Salmonella Enteritidis PT4 578 anaerobically cultivated in TSB at 37 °C in the presence or absence of C12-HSL.

(A) PCA of the logarithm of normalized TIC values of each triplicate of protein profile in the absence (control) and presence of C12-HSL. (B) Heatmap of the logarithm of normalized mean of TIC values of the triplicates for each identified protein. Each row corresponds to a unique protein, and each column the mean of the triplicate values. (C) Dendrogram of the logarithm normalized mean of TIC values of the triplicates for each identified protein. The height of the arms is proportional to the difference in the abundance profile of the proteins.

Fig 3. Number of differentially abundant proteins grouped according to the process on Gene Ontology (GO) annotations (European Bioinformatics Institute).

The Y-axis represents the number of differentially abundant proteins: above zero the number of proteins in which the abundance increased in the presence of C12-HSL compared to the control and below zero represents the proteins in which the abundance decreased in the presence of C12-HSL.

Fig 4. Proteins related to transcription process and “regulation of transcription, DNA-templated” function of Salmonella Enteritidis PT4 578, anaerobically cultivated in TSB at 37 °C in the presence or absence of C12-HSL.

(A) The network of interactions among the proteins of the transcription process and “regulation of transcription, DNA-templated” function and (B) the logarithm in base two of fold changed of the proteins that were identified at all times and in at least one of the treatments.

Fig 5. Identified proteins related to the oxidation-reduction process and quantification of free cellular thiol in Salmonella Enteritidis PT4 578 anaerobically cultivated in TSB at 37 °C in the presence or absence of C12-HSL.

(A) The network of interactions among the proteins of oxidation-reduction process and (B) the logarithm in base two of fold changed of the proteins that were identified at all time points and in at least one of the treatments, as well as (C) the quantification of free cellular thiol in the absence (control) and presence of C12-HSL.

Fig 6. Global response of Salmonella to the presence of C12-HSL.

The results related to a conclusion.