Virulence factors ofStaphylococcus aureusinduce Erk-MAP kinase activation and c-Fos expression in S9 and 16HBE14o- human airway epithelial cells

Abstract

Part of the innate defense of bronchial epithelia against bacterial colonization is regulated secretion of salt, water, and mucus as well as defensins and cytokines involving MAP kinase activation and alterations in early gene expression. We tested two different types of immortalized human airway epithelial cells (S9, 16HBE14o-) for activation of Erk-type MAP kinases and for expression of c-Fos on treatment with Staphylococcus aureus culture supernatants from the stationary growth phase [optical density (OD)(540 nm) = 10] or with recombinant S. aureus hemolysins A and B (Hla, Hlb). OD10 supernatants activated Erk-type MAP kinases and c-Fos expression in a concentration-dependent manner. Hla induced Erk-type kinase phosphorylation in S9 but not in 16HBE14o- cells. Hlb induced Erk activation in either cell type. Basal and stimulated levels of Erk-type MAP kinase phosphorylation were sensitive to the Mek1 inhibitor PD-98059, indicating that the bacterial products activated the entire signaling cascade that coregulates IL-8 induction and secretion. While c-Fos expression was enhanced by OD10 supernatants, Hla, and Hlb in S9 cells, 16HBE14o- cells responded to OD10 supernatant and Hlb but not to Hla. In S9 cells, PD-98059 suppressed c-Fos upregulation by OD10 supernatant, Hla, or Hlb, indicating that c-Fos expression requires activation of Erk-type MAP kinases. In 16HBE14o- cells, however, c-Fos expression by OD10 supernatant was sensitive to PD-98059, while that induced by Hlb was not. This indicates that ingredients of OD10 supernatants other than Hla or Hlb are activating Erk-type MAP kinases in 16HBE14o- cells and that other intracellular signaling systems apart from Erk-type MAP kinases contribute to Hlb-mediated regulation of c-Fos. Thus interaction of bacterial factors with airway epithelial cells may be highly cell type specific.

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

“…Lower concentrations of rHla may induce pore formation as well, but the physiological responses are much less pronounced and can hardly be detected. As we have previously shown, treatment of cells with 200 ng/ml rHla does not induce MAP kinase activation (4) or intracellular calcium accumulation (14).…”

“…These functional changes are supposed to represent defensive actions of the epithelial cells against the bacteria (32). On the other hand, exposure of epithelial cells to Hla may induce erratic intracellular signaling (4,19,25,45) in pathways mediating actin reorganization as well as loosening of cellmatrix adhesions (19), resulting in profound changes in cell shape (22,49) and in the formation of paracellular gaps in the previously confluent cell layer (19). In an in vivo situation, such cellular malfunctions may serve the bacteria to sustain their presence on host epithelial tissues and to establish infections.…”

“…2B). We conclude that disturbances of ion gradients across the plasma membrane and depolarization of the membrane potential are important inducers of defensive actions of these cells, as the activation of p38 MAP kinase is essential in the induction of IL-8 and IL-6 release (3,4,43).…”

“…Recombinant ␣-toxin (rHla) was prepared as described previously (4,14). The purity of rHla was assessed by SDS-PAGE after staining the gels with Coomassie brilliant blue.…”

“…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.…”