Human targets of Pseudomonas aeruginosa pyocyanin

Abstract

Pseudomonas aeruginosa produces copious amounts of the redoxactive tricyclic compound pyocyanin that kills competing microbes and mammalian cells, especially during cystic fibrosis lung infection. Cross-phylum susceptibility to pyocyanin suggests the existence of evolutionarily conserved physiological targets. We screened a Saccharomyces cerevisiae deletion library to identify presumptive pyocyanin targets with the expectation that similar targets would be conserved in humans. Fifty S. cerevisiae targets were provisionally identified, of which 60% have orthologous human counterparts. These targets encompassed major cellular pathways involved in the cell cycle, electron transport and respiration, epidermal cell growth, protein sorting, vesicle transport, and the vacuolar ATPase. Using cultured human lung epithelial cells, we showed that pyocyanin-mediated reactive oxygen intermediates inactivate human vacuolar ATPase, supporting the validity of the yeast screen. We discuss how the inactivation of VATPase may negatively impact the lung function of cystic fibrosis patients.T he most common pathogen during infection of the cystic fibrosis (CF) airways is Pseudomonas aeruginosa, which is also important in burn related sepsis, various nosocomial infections, and HIV-related illness (1, 2). The extensive host range and complex pathophysiology of infections associated with P. aeruginosa is due to its ability to produce a large repertoire of virulence determinants (1, 2). In addition to delivering proteins into eukaryotic cells by a type III secretion system to disrupt the host immune response and cause cytoskeletal reorganization (3), P. aeruginosa also produces a large number of toxic exoproducts, including proteases, rhamnolipids, and pyocyanin (PCN) (1, 2).PCN, a blue redox-active secondary metabolite, is a member of a large family of tricyclic compounds known as phenazines (4). Mutational and biochemical analyses have identified two groups of gene products required for PCN biosynthesis. First, the MvfR transcription factor is required to activate the phnAB genes (5-7). The phnA-B gene products synthesize quinolone that regulate the phzRABCDEFG operons 1 and 2, the structural genes for phenazine synthesis (8).Previous investigations have suggested that PCN contributes to the ability of P. aeruginosa to persist in the lungs of CF patients (9). In addition, in vitro studies have shown that PCN interferes with a myriad of mammalian cell functions. These include cell respiration, ciliary beating, epidermal cell growth (9, 10), calcium homeostasis (11), prostacyclin release from lung endothelial cells (12), apoptosis in neutrophils (13), release of interleukin-2 (limits growth of T lymphocytes), secretion of Ig by B-lymphocytes (14), and imbalance of protease-antiprotease activity in the airways of CF patients (15). However, the precise molecular mechanism underlying PCN-mediated pathology is unknown.PCN also inhibits microbial growth by initiating a redox cascade that can occur nonenzymatically via NADH or NADPH (16). Sev...

The power of chemical genomics to study the link between endomembrane system components and the gravitropic response

“…Finally, the chemical 5271050 is related to the phenazine family of chemicals, including pyocyanin, which is a secreted product of Pseudomonas aureginosa and contributes to virulence in cystic fibrosis patients (43). A yeast deletion strain library was screened on pyocyanin, and a number of strains defective for protein sorting, vesicle trafficking, and V-ATPase-related genes were shown to be hypersensitive to pyocyanin (43).…”

“…A yeast deletion strain library was screened on pyocyanin, and a number of strains defective for protein sorting, vesicle trafficking, and V-ATPase-related genes were shown to be hypersensitive to pyocyanin (43). This suggests possible targets in plants.…”

Calcium-Induced Virulence Factors Associated with the Extracellular Matrix of MucoidPseudomonas aeruginosaBiofilms

“…Therefore, it is likely that the in vivo Ca 2ϩ concentration in pulmonary fluid of CF patients is sufficient for induction of the phenazine biosynthetic genes. Pyocyanin is a redox cycling compound that has a number of effects on host cells, including an effect on the vacuolar ATPase (54). Among other functions, the vacuolar ATPase is important for maintaining cytosolic Ca 2ϩ levels in Saccharomyces cerevisiae (16).…”

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“…These compounds are known for their toxicity in other bacteria (Baron and Rowe, 1981) and in eukaryotes ranging from nematodes to humans (Ran et al, 2003;Lau et al, 2004;Mavrodi et al, 2006;Morales et al, 2010;Morales et al, 2013).…”