Pseudomonas aeruginosa pyocyanin modulates mucin glycosylation with sialyl-Lewisx to increase binding to airway epithelial cells

Abstract

Cystic fibrosis (CF) patients battle life-long pulmonary infections with the respiratory pathogen Pseudomonas aeruginosa (PA). An overabundance of mucus in CF airways provides a favorable niche for PA growth. When compared to that of non-CF individuals, mucus of CF airways is enriched in sialyl-Lewisx, a preferred binding receptor for PA. Notably, the levels of sialyl-Lewisx directly correlate with infection severity in CF patients. However, the mechanism by which PA causes increased sialylation remains uncharacterized. In this study, we examined the ability of PA virulence factors to modulate sialyl-Lewisx modification in airway mucins. We found pyocyanin (PCN) to be a potent inducer of sialyl-Lewisx in both mouse airways and in primary and immortalized CF and non-CF human airway epithelial cells. PCN increased the expression of C2/4GnT and ST3Gal-IV, two of the glycosyltransferases responsible for the stepwise biosynthesis of sialyl-Lewisx, through a TNF-α-mediated phosphoinositol-specific phospholipase C (PI-PLC) dependent pathway. Furthermore, PA bound more efficiently to airway epithelial cells pre-exposed to PCN through a flagellar cap-dependent manner. Importantly, antibodies against sialyl-Lewisx and anti-TNF-α attenuated PA binding. These results indicate that PCN secretes PCN to induce a favorable environment for chronic colonization of CF lungs by increasing the glycosylation of airway mucins with sialyl-Lewisx.

The Interaction between Respiratory Pathogens and Mucus

“…They are located on mucins and the surface of leukocytes, where they are involved in immune cell recruitment. There is evidence that PA can increase the Sialyl-Lewis X content on mucins via pyocyanin, a redox-active tricyclic toxin secreted by PA, thereby making the host environment more favorable for attachment and colonization (Jeffries et al, 2015). Pyocyanin has also been shown to stimulate the release of pro-inflammatory cytokines and MUC5AC and MUC2, which was mediated by the epidermal growth factor receptor (EGFR) pathway (Rada et al, 2011).…”

Role of Cytokine-Induced Glycosylation Changes in Regulating Cell Interactions and Cell Signaling in Inflammatory Diseases and Cancer

“…This increased adhesion was shown to be both dependent of the presence of sialic acid and of a functional FliD flagellar cap protein, a sLe x -binding protein [31]. Interestingly, a recent study suggests that the P. aeruginosa toxin pyocyanin itself is able to modulate sLe x expression on mucins in NCI-H292 cells, leading to an increased binding of P. aeruginosa [51]. Since the sialylation and sLe x expression of bronchial mucins secreted by patients suffering from CF or chronic bronchitis is related to the severity of airway infection [29], signaling pathways and molecular mechanisms responsible for TNF-induced over-expression of sLe x in human bronchial mucosa could be of critical importance to understand the links between sustained inflammation and P. aeruginosa infection observed in these lung pathologies.…”

FOXA2 depletion leads to mucus hypersecretion in canine airways with respiratory diseases

“…Further detailed studies indicate that P. aeruginosa and pyocyanin downregulate the expression of FOXA2, which is accompanied by excessive expression and secretion of MUC5AC and MUC5B mucins (Caldwell et al, 2009;Hao et al, 2012Hao et al, , 2013Jeffries et al, 2016;Xu et al, 2013). Further detailed studies indicate that P. aeruginosa and pyocyanin downregulate the expression of FOXA2, which is accompanied by excessive expression and secretion of MUC5AC and MUC5B mucins (Caldwell et al, 2009;Hao et al, 2012Hao et al, , 2013Jeffries et al, 2016;Xu et al, 2013).…”

“…Paraffin-embedded canine lung tissue sections were stained with PAS reagents (395B, Sigma-Aldrich) or by IHC as we have previously published (Caldwell et al, 2009;Hao et al, 2012;Hao et al, 2013;Jeffries et al, 2016;Xu et al, 2013). For IHC, lung sections were probed with primary antibodies and detected using ABC kit (PK-6101, PK-105, Vector Laboratories) according to the protocol supplied by the manufacturers.…”

“…;Jeffries et al, 2016;Lau, Ran, Kong, Hassett, & Mavrodi, 2004;Xu et al, 2013) induces GCHM, mucus hypersecretion, fibrosis, and emphysema in mouse airways and in both immortalised and primary human airway epithelial cells by activating STAT6-SPDEF and EGFR-AKT/ERK1/2 signalling pathways that antagonise FOXA2. To determine the mechanism by which P. aeruginosa depletes FOXA2 expression, induces GCHM, and mucus hypersecretion in canine lungs, BACA cells, an immortalised canine airway carcinoma cell line, were infected with the wild-type P. aeruginosa strain PAO1 at multiplicity of infection (MOI) of 1:1 and 1:10 for 6, 12, and 24 hr.…”