Cytotoxic Lung Endothelial Amyloids

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PROJECT SUMMARY/ABSTRACT Pulmonary endothelial barrier integrity is vital for efficient gas exchange and to prevent the hematogenous dissemination of respiratory pathogens. Pseudomonas aeruginosa, a predominant agent of nosocomial pneumonia, breaches the endothelial barrier by inducing junctional complex disruption leading to alveolar flooding and hypoxemia. Here, we propose to study a previously unidentified, novel disease process defined by the interaction of the common nosocomial pathogen, P. aeruginosa, with the host endothelium. P. aeruginosa utilizes a Type III Secretion System (T3SS) to inject cytotoxic exoenzymes directly into the host cell. Exoenzyme Y (ExoY), a T3SS effector expressed by ~90% of clinical P. aeruginosa strains, causes marked increases in cytosolic cGMP, cUMP, and cAMP. The increase in cAMP facilitates the phosphorylation of an endothelial tau, which dissociates from microtubules, followed by microtubule collapse, interendothelial gap formation, and increased permeability. Hyperphosphorylated tau then coalesces into cytotoxic tau oligomers that are released into the extracellular milieu. These cytotoxic tau oligomers are transmissibleand self-propagating amyloid prions. The respective roles of cGMP and cUMP are currently unknown, although preliminary data establishes both a temporal and spatial correlation between the increase in ExoY-generated cUMP and oligomeric tau release from PMVECs. The cUMP signal in the bulk cytosol closely coincides with both the reduction of cytosolic tau and the concomitant increase of extracellular tau. Thus, an ExoY-instigated increase in cUMP is implicated in the export of cytotoxic tau. Furthermore, within the context of neurodegenerative disease, the degradation and recycling pathway of autophagy is often pathologically constrained thereby preventing the breakdown of excess or dysregulated amyloids. Following the inhibition of autophagic flux, dysregulated amyloids accumulate and oligomerize. Clearance of nascent tau oligomers is often facilitated via exosomal and/or secretion mediated export. Concordant with our observations, ExoY has recently been implicated in the suppression of the innate immune response via the inhibition of transforming growth factor-β-activating kinase 1 (TAK1) which has also been reported to obstruct autophagy. Taken together, ExoY suppression of TAK1 in PMVECs may contribute to the inhibition of autophagy and the production of infection-induced tau prions. Therefore, this proposal tests the hypothesis that ExoY-induced cUMP contributes to the suppression of autophagic flux thereby promoting the generation and release of cytotoxic tau that transmissibly disrupts interendothelial junctions and promotes perm eability. .