Abstract
Airway epithelial cells reduce cytosolic ATP content in response to treatment with S. aureus alpha-toxin (hemolysin A, Hla). This study was undertaken to investigate whether this is due to attenuated ATP generation or to release of ATP from the cytosol and extracellular ATP degradation by ecto-enzymes. Exposure of cells to rHla did result in mitochondrial calcium uptake and a moderate decline in mitochondrial membrane potential, indicating that ATP regeneration may have been attenuated. In addition, ATP may have left the cells through transmembrane pores formed by the toxin or through endogenous release channels (e.g., pannexins) activated by cellular stress imposed on the cells by toxin exposure. Exposure of cells to an alpha-toxin mutant (H35L), which attaches to the host cell membrane but does not form transmembrane pores, did not induce ATP release from the cells. The Hla-mediated ATP-release was completely blocked by IB201, a cyclodextrin-inhibitor of the alpha-toxin pore, but was not at all affected by inhibitors of pannexin channels. These results indicate that, while exposure of cells to rHla may somewhat reduce ATP production and cellular ATP content, a portion of the remaining ATP is released to the extracellular space and degraded by ecto-enzymes. The release of ATP from the cells may occur directly through the transmembrane pores formed by alpha-toxin.
Highlights
Many strains of Staphylococcus aureus produce virulence factors that are secreted to the external medium and mediate pathogenicity [1,2]
Hla is secreted from the cells as a 33 kDa monomer that attaches to the outer surface of eukaryotic host cells
Hla may associate with eukaryotic plasma membranes in a specific manner with half-maximal binding at 1–2 nmol/L (30–60 ng/mL) [6]
Summary
Many strains of Staphylococcus aureus produce virulence factors that are secreted to the external medium and mediate pathogenicity [1,2]. S. aureus strains is the pore-forming alpha-toxin (hemolysin A, Hla) [3,4]. Hla is secreted from the cells as a 33 kDa monomer that attaches to the outer surface of eukaryotic host cells. Hla may associate with eukaryotic plasma membranes in a specific manner with half-maximal binding at 1–2 nmol/L (30–60 ng/mL) [6]. Areas of specific lipid composition may be involved in Hla attachment to the host cell plasma membrane [8]. Non-specific binding, occurs at concentrations at >1 μmol/L (33 μg/mL) [6], which may result in cell disruption (e.g., hemolysis) by disturbing the lipid barrier
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