Staphylococcus aureus Hemolysin A Disrupts Cell–Matrix Adhesions in Human Airway Epithelial Cells

Abstract

Treatment of primary or immortalized human airway epithelial cells (16HBE14o-, S9) or alveolar cancer cells (A549) with recombinant hemolysin A (rHla), a major virulence-associated factor of Staphylococcus aureus, induces alterations in cell shape and formation of paracellular gaps in the cell layer. Semiquantitative Western blotting using extracts of freshly isolated airway tissue (nasal epithelium) or 16HBE14o- model cells revealed that phosphorylation levels of focal adhesion kinase (Fak) and paxillin were altered upon treatment of tissue or cells with rHla. Immune fluorescence analyses showed that rHla treatment of 16HBE14o- cells results in losses of vinculin and paxillin from focal contacts and a net reduction in the number of focal contacts. The actin cytoskeleton was strongly remodeled. We concluded that treatment of cells with rHla activates Fak signaling, which accelerates focal contact turnover and prevents newly formed focal contacts (focal complexes) from maturation to focal adhesions. The inability of rHla-treated cells to form stable focal adhesions may be one factor that contributes to gap formation in the cell layer. In vivo, such changes may disturb the defensive barrier function of the airway epithelium and may facilitate lung infections by S. aureus.

Validation of Normal Human Bronchial Epithelial Cells as a Model for Influenza A Infections in Human Distal Trachea

“…2I). These findings demonstrated that, at a minimum, the basal plasma membranes of some NHBE cells were forming focal complexes with extracellular matrix or membrane substance (Berrier and Yamada, 2007;Hermann et al 2014).…”

Staphylococcus aureusα-toxin-mediated cation entry depolarizes membrane potential and activates p38 MAP kinase in airway epithelial cells

“…Treatment of eukaryotic cells with Hla elicits various cell physiological effects ranging from apoptotic or necrotic cell death (16) or survival signaling (24,43) to changes in cell-cell or cell-matrix adhesion and actin cytoskeleton reorganization (19,60). Studies using cultured airway epithelial cells revealed that they respond to Hla treatment with ADAM10-mediated cleavage of cadherin (25).…”

“…Studies using cultured airway epithelial cells revealed that they respond to Hla treatment with ADAM10-mediated cleavage of cadherin (25). We have recently shown that treatment of airway epithelial cells with recombinant Hla (rHla) results in accelerated turnover of focal adhesion complexes resulting in a net loss of peripheral focal contacts (19). Concomittant restructuring of the actin cytoskeleton results in the inability of cells to stabilize their cell shapes and in the appearance of paracellular gaps in the cell layer (19).…”

“…We have recently shown that treatment of airway epithelial cells with recombinant Hla (rHla) results in accelerated turnover of focal adhesion complexes resulting in a net loss of peripheral focal contacts (19). Concomittant restructuring of the actin cytoskeleton results in the inability of cells to stabilize their cell shapes and in the appearance of paracellular gaps in the cell layer (19). Such changes correspond to epithelial injury and loss of epithelial barrier function as observed during S. aureus infections in the airways (5).…”

“…Hla treatment of airway epithelial cells results in activation of ADAM10 (60), which may affect several subsequent signaling modules by as yet unknown links in the transduction pathways including proinflammatory pathways (13). However, ADAM10 activation depends on the presence of extracellular calcium (25) and on calcium influx into the cytosol (23 (4,19,43), but the modes of activation are still elusive. The aim of this study was to elucidate whether mono-or divalent cations can pass through the Hla pores in otherwise intact epithelial cells and whether Hla-mediated changes in the cytosolic concentrations of mono-or divalent cations are sufficient to activate intracellular signaling pathways affecting the above mentioned cellular processes.…”