Pseudomonas aeruginosa lung infections are life threatening and constitute a large health-care burden. Modulation of host immune responses is one paradigm used by pathogens to initiate and prolong infections. Preliminary data demonstrate that P. aeruginosa can degrade the potent immune signaling molecule sphingosine-1-phosphate (S1P) and its precursor sphingosine. S1P degradation would allow P. aeruginosa to alter the host immune response, which could have profound effects on progression of infection and the pathogenesis of disease. Described in this proposal is a previously uncharacterized transcription factor that regulates P. aeruginosa gene expression in response to S1P and sphingosine and is required for their degradation. Deletion of this transcription factor reduces P. aeruginosa virulence in vivo, highlighting the importance of this pathway during infection. Characterization of the detection and metabolism of sphingosine by P. aeruginosa will be a concrete contribution to understanding of sphingosine-related compounds in bacterial biology and pathogenesis. The long term goal is to understand the role of P. aeruginosa S1P and sphingosine metabolism during the course of lung infection. In pursuit of this long-term goal, the overall objectives of this application are to characterize the recognition and degradation system used by P. aeruginosa to respond to S1P and determine the impact of S1P degradation on P. aeruginosa virulence in the lung. The central hypothesis is that P. aeruginosa detection of S1P induces genes necessary for S1P degradation, which promotes virulence. The central hypothesis will be tested by pursuing three specific aims: (1) Characterize S1P and sphingosine-dependent transcription in P. aeruginosa~ (2) Identify the components of the S1P degradation pathway~ and (3) Test the impact of S1P degradation on P. aeruginosa infection. The goals of Aim 1 are to determine the ligand and promoter-binding specificities of the sphingosine-responsive transcription factor by promoter mapping, DNA binding, and reporter assays, and determine its contribution to the sphingosine regulon using microarray analysis. The goal of Aim 2 is to identify the P. aeruginosa S1P phosphatase and the enzymes that degrade sphingosine using genetic screens, metabolite tracking, and in vitro enzymatic assays. Finally, the goal of Aim 3 is to test the hypothesis that the reduced virulence of the transcription factor mutant is due to lack of S1P degradation, versus regulation of genes unrelated to degradation, using a mouse model of lung infection established in the applicant's lab. This proposal is innovative because it will advance understanding of a previously uncharacterized sphingosine detection and metabolic pathway, which may be used by P. aeruginosa to detect the host and interfere with a potent host signaling pathway. The proposed research is significant because it will expand understanding of the metabolic links between bacterial pathogens and the host, perhaps leading to novel antimicrobial or anti-virulence therapies.

Public Health Relevance

The research proposed here is relevant to human health because bacterial detection and modulation of host immune responses is used by pathogens to cause infections, and S1P degradation represents a novel mechanism for P. aeruginosa virulence. This proposal is relevant to the NIH mission both in terms of developing fundamental knowledge about pathogens to benefit human health and because S1P degradation by bacterial pathogens may represent therapeutic targets to treat infections.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI103003-05
Application #
9170956
Study Section
Bacterial Pathogenesis Study Section (BACP)
Program Officer
Lu, Kristina
Project Start
2012-11-15
Project End
2017-10-31
Budget Start
2016-11-01
Budget End
2017-10-31
Support Year
5
Fiscal Year
2017
Total Cost
Indirect Cost
Name
University of Vermont & St Agric College
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
066811191
City
Burlington
State
VT
Country
United States
Zip Code
05405
Willsey, Graham G; Ventrone, Sebastian; Schutz, Kristin C et al. (2018) Pulmonary Surfactant Promotes Virulence Gene Expression and Biofilm Formation in Klebsiella pneumoniae. Infect Immun 86:
Meadows, Jamie A; Wargo, Matthew J (2018) Transcriptional Regulation of Carnitine Catabolism in Pseudomonas aeruginosa by CdhR. mSphere 3:
English, Erika L; Schutz, Kristin C; Willsey, Graham G et al. (2018) Transcriptional Responses of Pseudomonas aeruginosa to Potable Water and Freshwater. Appl Environ Microbiol 84:
Meadows, Jamie A; Willsey, Graham G; Wargo, Matthew J (2018) Differential requirements for processing and transport of short-chain versus long-chain O-acylcarnitines in Pseudomonas aeruginosa. Microbiology 164:635-645
Burgess, Edward J; Hoyt, Laura R; Randall, Matthew J et al. (2018) Bacterial Lipoproteins Constitute the TLR2-Stimulating Activity of Serum Amyloid A. J Immunol 201:2377-2384
Demirdjian, Sally; Schutz, Kristin; Wargo, Matthew J et al. (2017) The effect of loss of O-antigen ligase on phagocytic susceptibility of motile and non-motile Pseudomonas aeruginosa. Mol Immunol 92:106-115
Nock, Adam M; Wargo, Matthew J (2016) Choline Catabolism in Burkholderia thailandensis Is Regulated by Multiple Glutamine Amidotransferase 1-Containing AraC Family Transcriptional Regulators. J Bacteriol 198:2503-14
Ubags, Niki D J; Burg, Elianne; Antkowiak, Maryellen et al. (2016) A Comparative Study of Lung Host Defense in Murine Obesity Models. Insights into Neutrophil Function. Am J Respir Cell Mol Biol 55:188-200
Ubags, Niki D J; Stapleton, Renee D; Vernooy, Juanita H J et al. (2016) Hyperleptinemia is associated with impaired pulmonary host defense. JCI Insight 1:
Willsey, Graham G; Wargo, Matthew J (2016) Sarcosine Catabolism in Pseudomonas aeruginosa Is Transcriptionally Regulated by SouR. J Bacteriol 198:301-10

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