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Biofilm Formation Mechanisms of Pseudomonas aeruginosa Predicted via Genome-Scale Kinetic Models of Bacterial Metabolism.

Vital-Lopez FG, Reifman J, Wallqvist A - PLoS Comput. Biol. (2015)

Bottom Line: Our analysis suggests that the synthesis of important biofilm-related molecules, such as the quorum-sensing molecule Pseudomonas quinolone signal and the exopolysaccharide Psl, is regulated not only through the expression of genes in their own synthesis pathway, but also through the biofilm-specific expression of genes in pathways competing for precursors to these molecules.Alternative to a previous hypothesis that this biofilm reduction is caused by a decrease in energy production, we proposed that the dysregulation of the synthesis of secondary metabolites derived from anthranilate and chorismate is what impaired the biofilms of these mutants.Notably, these insights generated through our kinetic model-based approach are not accessible from previous constraint-based model analyses of P. aeruginosa biofilm metabolism.

View Article: PubMed Central - PubMed

Affiliation: Department of Defense Biotechnology High Performance Computing Software Applications Institute, Telemedicine and Advanced Technology Research Center, U.S. Army Medical Research and Materiel Command, Fort Detrick, Maryland, United States of America.

ABSTRACT
A hallmark of Pseudomonas aeruginosa is its ability to establish biofilm-based infections that are difficult to eradicate. Biofilms are less susceptible to host inflammatory and immune responses and have higher antibiotic tolerance than free-living planktonic cells. Developing treatments against biofilms requires an understanding of bacterial biofilm-specific physiological traits. Research efforts have started to elucidate the intricate mechanisms underlying biofilm development. However, many aspects of these mechanisms are still poorly understood. Here, we addressed questions regarding biofilm metabolism using a genome-scale kinetic model of the P. aeruginosa metabolic network and gene expression profiles. Specifically, we computed metabolite concentration differences between known mutants with altered biofilm formation and the wild-type strain to predict drug targets against P. aeruginosa biofilms. We also simulated the altered metabolism driven by gene expression changes between biofilm and stationary growth-phase planktonic cultures. Our analysis suggests that the synthesis of important biofilm-related molecules, such as the quorum-sensing molecule Pseudomonas quinolone signal and the exopolysaccharide Psl, is regulated not only through the expression of genes in their own synthesis pathway, but also through the biofilm-specific expression of genes in pathways competing for precursors to these molecules. Finally, we investigated why mutants defective in anthranilate degradation have an impaired ability to form biofilms. Alternative to a previous hypothesis that this biofilm reduction is caused by a decrease in energy production, we proposed that the dysregulation of the synthesis of secondary metabolites derived from anthranilate and chorismate is what impaired the biofilms of these mutants. Notably, these insights generated through our kinetic model-based approach are not accessible from previous constraint-based model analyses of P. aeruginosa biofilm metabolism. Our simulation results showed that plausible, non-intuitive explanations of difficult-to-interpret experimental observations could be generated by integrating genome-scale kinetic models with gene expression profiles.

No MeSH data available.


Related in: MedlinePlus

Expression of pqs, psl, and pel operons as a function of biofilm age.(A) Correlation coefficient of the expression intensity of the genes in the pqs, psl, and pel operons and biofilm age (h). (B) Expression of one gene from each operon as a function of biofilm age. Gene expressions are shown in log2 scale. Each dot corresponds to one condition in the dataset of gene expression for P. aeruginosa PAO1 biofilms. The circles correspond to the data obtained by Costaglioli et al. [11]. AU, arbitrary units; h, hours.
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pcbi.1004452.g002: Expression of pqs, psl, and pel operons as a function of biofilm age.(A) Correlation coefficient of the expression intensity of the genes in the pqs, psl, and pel operons and biofilm age (h). (B) Expression of one gene from each operon as a function of biofilm age. Gene expressions are shown in log2 scale. Each dot corresponds to one condition in the dataset of gene expression for P. aeruginosa PAO1 biofilms. The circles correspond to the data obtained by Costaglioli et al. [11]. AU, arbitrary units; h, hours.

Mentions: We predicted that the synthesis of two biofilm-related molecules would have a considerable flux change without significant expression changes of the genes associated with the corresponding enzymatic reactions. The production of PQS was reduced, while the synthesis of the polysaccharide Psl was increased. PQS is a quorum-sensing signal that regulates the production of virulence factors and biofilm development in a complex regulatory network with the N-acylhomoserine lactone-dependent quorum-sensing systems las and rhl [30]. Diggle et al. [31] reported increased biofilm formation in P. aeruginosa cultures treated with exogenous PQS after 72 h. Similarly, Guo et al. [32] reported that PQS promoted biofilm formation, but in their experiments the enhancement occurred within the first hour of incubation, and the cultures with and without exogenous PQS had similar biofilm formation rates from 1 to 24 h. This observation, coupled with the observation that the genes involved in PQS synthesis were not up-regulated in the biofilm, suggests that PQS may be more important at the early stages of biofilm formation and that its synthesis is down-regulated as the biofilm matures. Moreover, our simulation results indicated that further repression of PQS synthesis was achieved by increasing anthranilate degradation. We investigated whether PQS is indeed down-regulated as the biofilm matures, rather than as a consequence of the specific experimental condition in the study by Costaglioli et al. [11]. To this end, we analyzed the expression of the PQS synthesis genes (pqsABCDH) in all datasets available in the Gene Expression Omnibus (GEO) database [33] for the wild-type PAO1 strain of P. aeruginosa growing in biofilm (see Materials and Methods for details on processing the gene expression data). We found that there was a significant negative correlation between the expression of the PQS synthesis genes and the biofilm age (Fig 2).


Biofilm Formation Mechanisms of Pseudomonas aeruginosa Predicted via Genome-Scale Kinetic Models of Bacterial Metabolism.

Vital-Lopez FG, Reifman J, Wallqvist A - PLoS Comput. Biol. (2015)

Expression of pqs, psl, and pel operons as a function of biofilm age.(A) Correlation coefficient of the expression intensity of the genes in the pqs, psl, and pel operons and biofilm age (h). (B) Expression of one gene from each operon as a function of biofilm age. Gene expressions are shown in log2 scale. Each dot corresponds to one condition in the dataset of gene expression for P. aeruginosa PAO1 biofilms. The circles correspond to the data obtained by Costaglioli et al. [11]. AU, arbitrary units; h, hours.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4592021&req=5

pcbi.1004452.g002: Expression of pqs, psl, and pel operons as a function of biofilm age.(A) Correlation coefficient of the expression intensity of the genes in the pqs, psl, and pel operons and biofilm age (h). (B) Expression of one gene from each operon as a function of biofilm age. Gene expressions are shown in log2 scale. Each dot corresponds to one condition in the dataset of gene expression for P. aeruginosa PAO1 biofilms. The circles correspond to the data obtained by Costaglioli et al. [11]. AU, arbitrary units; h, hours.
Mentions: We predicted that the synthesis of two biofilm-related molecules would have a considerable flux change without significant expression changes of the genes associated with the corresponding enzymatic reactions. The production of PQS was reduced, while the synthesis of the polysaccharide Psl was increased. PQS is a quorum-sensing signal that regulates the production of virulence factors and biofilm development in a complex regulatory network with the N-acylhomoserine lactone-dependent quorum-sensing systems las and rhl [30]. Diggle et al. [31] reported increased biofilm formation in P. aeruginosa cultures treated with exogenous PQS after 72 h. Similarly, Guo et al. [32] reported that PQS promoted biofilm formation, but in their experiments the enhancement occurred within the first hour of incubation, and the cultures with and without exogenous PQS had similar biofilm formation rates from 1 to 24 h. This observation, coupled with the observation that the genes involved in PQS synthesis were not up-regulated in the biofilm, suggests that PQS may be more important at the early stages of biofilm formation and that its synthesis is down-regulated as the biofilm matures. Moreover, our simulation results indicated that further repression of PQS synthesis was achieved by increasing anthranilate degradation. We investigated whether PQS is indeed down-regulated as the biofilm matures, rather than as a consequence of the specific experimental condition in the study by Costaglioli et al. [11]. To this end, we analyzed the expression of the PQS synthesis genes (pqsABCDH) in all datasets available in the Gene Expression Omnibus (GEO) database [33] for the wild-type PAO1 strain of P. aeruginosa growing in biofilm (see Materials and Methods for details on processing the gene expression data). We found that there was a significant negative correlation between the expression of the PQS synthesis genes and the biofilm age (Fig 2).

Bottom Line: Our analysis suggests that the synthesis of important biofilm-related molecules, such as the quorum-sensing molecule Pseudomonas quinolone signal and the exopolysaccharide Psl, is regulated not only through the expression of genes in their own synthesis pathway, but also through the biofilm-specific expression of genes in pathways competing for precursors to these molecules.Alternative to a previous hypothesis that this biofilm reduction is caused by a decrease in energy production, we proposed that the dysregulation of the synthesis of secondary metabolites derived from anthranilate and chorismate is what impaired the biofilms of these mutants.Notably, these insights generated through our kinetic model-based approach are not accessible from previous constraint-based model analyses of P. aeruginosa biofilm metabolism.

View Article: PubMed Central - PubMed

Affiliation: Department of Defense Biotechnology High Performance Computing Software Applications Institute, Telemedicine and Advanced Technology Research Center, U.S. Army Medical Research and Materiel Command, Fort Detrick, Maryland, United States of America.

ABSTRACT
A hallmark of Pseudomonas aeruginosa is its ability to establish biofilm-based infections that are difficult to eradicate. Biofilms are less susceptible to host inflammatory and immune responses and have higher antibiotic tolerance than free-living planktonic cells. Developing treatments against biofilms requires an understanding of bacterial biofilm-specific physiological traits. Research efforts have started to elucidate the intricate mechanisms underlying biofilm development. However, many aspects of these mechanisms are still poorly understood. Here, we addressed questions regarding biofilm metabolism using a genome-scale kinetic model of the P. aeruginosa metabolic network and gene expression profiles. Specifically, we computed metabolite concentration differences between known mutants with altered biofilm formation and the wild-type strain to predict drug targets against P. aeruginosa biofilms. We also simulated the altered metabolism driven by gene expression changes between biofilm and stationary growth-phase planktonic cultures. Our analysis suggests that the synthesis of important biofilm-related molecules, such as the quorum-sensing molecule Pseudomonas quinolone signal and the exopolysaccharide Psl, is regulated not only through the expression of genes in their own synthesis pathway, but also through the biofilm-specific expression of genes in pathways competing for precursors to these molecules. Finally, we investigated why mutants defective in anthranilate degradation have an impaired ability to form biofilms. Alternative to a previous hypothesis that this biofilm reduction is caused by a decrease in energy production, we proposed that the dysregulation of the synthesis of secondary metabolites derived from anthranilate and chorismate is what impaired the biofilms of these mutants. Notably, these insights generated through our kinetic model-based approach are not accessible from previous constraint-based model analyses of P. aeruginosa biofilm metabolism. Our simulation results showed that plausible, non-intuitive explanations of difficult-to-interpret experimental observations could be generated by integrating genome-scale kinetic models with gene expression profiles.

No MeSH data available.


Related in: MedlinePlus