Significance of Corneal Cell Invasion By Bacteria

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DESCRIPTION (provided by applicant): Corneal epithelial barrier function against microbes is generally thought to involve surface mucins, junctional barriers, and extracellular antimicrobials. Additionally, we have found that corneal epithelial cells can behave as "non-professional phagocytes", i.e. they can internalize bacteria and then suppress their viability within acidified lysosomes. The importance of this potential innate defense against microbes remains to be determined and is a goal of this project. The other goal is to determine how bacteria evade this defense. Pseudomonas aeruginosa is among the most common and the most devastating of corneal pathogens. The statistics are even worse for other epithelial lined body sites, where P. aeruginosa infection very often results in mortality. While often referred to as an extracellular pathogen, we have shown P. aeruginosa can thrive in corneal (and airway) epithelial cells, by evading acidified lysosomes and establishing replicative niches inside plasma membrane blebs. While lysosome evasion and "bleb-niche" formation both require the bacterial type III secretion system (T3SS), these capacities are separable. Lysosome evasion depends on the ADP-ribosylation (ADPr) activity of the T3SS effector ExoS, but not PopB, one of two T3SS translocators. In stark contrast, both ExoS and PopB are needed for P. aeruginosa to establish bleb-niches. The three aims of this project are: Aim 1. To study mechanisms by which corneal epithelial cells kill intracellular bacteria; specifically exploring the role of MyD88, which we hae found is involved. Aim 2: To determine how P. aeruginosa uses ExoS to evade acidified lysosomes in corneal epithelial cells. Aim 3. To explore how PopB and ExoS then allow intracellular P. aeruginosa to establish membrane bleb niches. Phagocytic activity of corneal epithelial cells and bacterial evasion of this defense are both novel concepts. Understanding the host-pathogen interactions that result in these very different outcomes, could lead to new strategies for managing infection.