Monday, December 30, 2024
12/30/2024
PROJECT SUMMARY/ABSTRACT The alveolar-capillary membrane facilitates efficient gas exchange while maintaining a restrictive permeability barrier. Pseudomonas aeruginosa infection disrupts the alveolar-capillary barrier leading to exudative edema and impaired oxygenation. P. aeruginosa utilizes a type III secretion system and its effectors to disrupt barrier integrity. In particular, the exoenzyme Y is introduced into lung endothelium, where it acquires nucleotidylyl cyclase activity and produces cGMP, cAMP, and cUMP. These cyclic nucleotide monophosphates activate protein kinase A resulting in endothelial tau phosphorylation, tau dissociation from microtubules, and microtubule breakdown, which collectively hinders repair following infection. Phosphorylated tau is released from endothelium as cytotoxic variants that contribute to lung injury. The signaling mechanisms used by exoenzyme Y to produce cytotoxic tau is incompletely understood, yet cUMP is produced at especially high concentrations and the cUMP signal parallels the generation of cytotoxic tau. Elevations in cUMP are sufficient to promote the production of cytotoxic tau variants. Our preliminary data demonstrate that the exoenzyme Y-induced cUMP signal also decreases endothelial nicotinamide adenine dinucleotide (NAD+) and increases nicotinamide, the product of NAD+ hydrolase activity, which may impair recovery following infection. Lung endothelium expresses sterile alpha and TIR motif containing 1 (SARM1), the only TIR (Toll/Interleukin-1 Receptor) domain protein in mammalian cells that possesses NAD+ hydrolase activity. Recent studies revealed a SARM1 bacterial homologue is directly activated by cUMP as an essential innate immune mechanism. While our studies illustrate an important role for cUMP in the endothelial cell response to infection, how exoenzyme Y generates the cUMP that leads to tau phosphorylation and production of cytotoxic tau variants, and how cUMP lowers NAD+ while hindering endothelial cell repair remains poorly understood. To address this knowledge gap in a rigorous way, this project tests the hypothesis that the P. aeruginosa exoenzyme Y generates cUMP, which contributes to the tau phosphorylation, microtubule breakdown, and SARM1-dependent NAD+ hydrolase activity that causes lung injury and hinders repair.
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