Bacterial pathogens often damage their host through the action of toxins. Bacterial toxins modify host targets, primarily proteins, through covalent and non-covalent mechanisms and have several structure-function organizations, including exotoxins and type III secreted cytotoxins. While exotoxins often act at a distance from the site of infection, type III cytotoxins are delivered directly into the intoxicated host cell by the bacterium.
The aims of this application have been to characterize the substrate recognition and intracellular trafficking of bacterial toxins. During the previous period, studies on the type III cytotoxins of Pseudomonas aeruginosa determined substrate recognition mechanisms of ExoS and ExoT and showed that intracellular trafficking within host cells increased the potency of ExoS for Ras and Rho GTPases. A translational project was also developed to detect initial PA infections in humans. This renewal will continue to characterize the intracellular trafficking and substrate recognition on ExoS and initiate structure-function studies on tetanus neurotoxin (TeNT). The proposed studies will characterize specific properties of ExoS and TeNT that define unique aspects of bacterial toxin action. Analyses utilize biochemical- and quantitative cell biological- approaches that allow corroborative analyses from the test tube to the cell. Studies on ExoS will characterize how intracellular trafficking within host cells increases the potency of ExoS and determine the mechanism that ADP-ribosylation inhibits signaling by small GTPases, like Ras and Rab5. Recent studies determined that gangliosides are the functional receptors of TeNT in neurons. This identification allows a fundamental question of neurotoxin pathology to be addressed, how does TeNT elicit spastic paralysis in the host? Studies on TeNT will characterize the intracellular trafficking of TeNT in neurons and determine how TeNT cleaves the SNARE protein, VAMP2. These studies define mechanistic properties of toxins that may lead to translational products for the detection, prevention, and intervention of bacterial infections.

Public Health Relevance

Bacterial toxins damage host cells by directly modifying intracellular targets, which may lead to cell death. This renewal will study how bacterial toxins traffic within host cells and recognize host proteins that are targeted for modification. Studies are proposed for ExoS, a toxin produced by Pseudomonas aeruginosa, which is responsible for many hospital- acquired infections and infections of compromised individuals including children with cystic fibrosis, and tetanus toxin, a neurotoxin that elicits spastic paralysis in humans. Characterization of these bacterial toxins will provide information on how bacteria damage the host and may lead to translational products to detect, prevent, and treat bacterial infections.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Host Interactions with Bacterial Pathogens Study Section (HIBP)
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Taylor, Christopher E,
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Medical College of Wisconsin
Schools of Medicine
United States
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Zuverink, Madison; Barbieri, Joseph T (2018) Protein Toxins That Utilize Gangliosides as Host Receptors. Prog Mol Biol Transl Sci 156:325-354
Zuverink, Madison; Barbieri, Joseph T (2017) Protein Structure Facilitates High-Resolution Immunological Mapping. Clin Vaccine Immunol 24:
Kroken, Abby R; Blum, Faith C; Zuverink, Madison et al. (2017) Entry of Botulinum Neurotoxin Subtypes A1 and A2 into Neurons. Infect Immun 85:
Chen, Chen; Barbieri, Joseph T (2017) When Escherichia coli doesn't fit the mold: A pertussis-like toxin with altered specificity. J Biol Chem 292:15159-15160
Chen, Sheng; Barbieri, Joseph T (2016) Solubility of the catalytic domains of Botulinum neurotoxin serotype E subtypes. Protein Expr Purif 118:18-24
Zuverink, Madison; Chen, Chen; Przedpelski, Amanda et al. (2015) A Heterologous Reporter Defines the Role of the Tetanus Toxin Interchain Disulfide in Light-Chain Translocation. Infect Immun 83:2714-24
Zuverink, Madison; Barbieri, Joseph T (2015) From GFP to ?-lactamase: advancing intact cell imaging for toxins and effectors. Pathog Dis 73:ftv097
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Blum, Faith C; Tepp, William H; Johnson, Eric A et al. (2014) Multiple domains of tetanus toxin direct entry into primary neurons. Traffic 15:1057-65

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