Date of Award

2023

Degree Type

Thesis

Degree Name

Master of Science (MS)

Department

Microbiology and Immunology

Abstract

Pseudomonas aeruginosa is a gram-negative environmental bacterium that uses its intrinsic and acquired antibiotic resistance to infect patients with compromised epithelium, such as cystic fibrosis and burn wounds, and corneal keratitis. In order to cause disease, most strains of P. aeruginosa use the type three secretion system (T3SS) to disrupt cellular signaling and integrity. However, P. aeruginosa may also secrete exotoxins after invading host cells, which prolongs host cell death and maintains an intracellular niche. Of the T3 secreted effector toxins, ExoS, specifically the ADP-ribosyltransferase activity of ExoS, was shown to delay the death of invaded corneal cells, but it is dispensable for maintaining intact intracellular niches when caspase-4 is deleted. This suggests ExoS interferes with the completion of caspase-4 mediated pyroptosis. While ExoS has no known substrates involved in caspase-4 mediated cell death, other groups have demonstrated many potential ExoS substrates remain uncategorized. We hypothesized that ExoS ADP-ribosylates a target in the caspase-4 inflammasome to delay corneal cell death, and developed methods to identify new ExoS substrates. As a consequence of investigating new tools to detect ADP-ribosylation, we were also able to develop a method to visualize T3SS-targeted cells simultaneously with intracellular P. aeruginosa. Using these methods, and inhibitors of bacterial invasion, we found that bacterial internalization is important for delivery of T3SS effectors into corneal epithelial cells. In sum, this work further illustrates the necessity of P. aeruginosa intracellular lifestyle.

Creative Commons License

Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License
This work is licensed under a Creative Commons Attribution-Noncommercial-No Derivative Works 3.0 License.

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