Pore formation by a two-component leukocidin from Staphylococcus aureus within the membrane of human polymorphonuclear leukocytes.
The effects of the Staphylococcus aureus leukocidin (PVL), a two-component non-hemolytic toxin, were investigated on the membrane permeability of human polymorphonuclear leukocytes (PMNs). In the absence of extracellular Ca2+, the fluorescence of ethidium bromide added to the extracellular medium increased after PVL application in a concentration-dependent manner and no variations in the free intracellular [Ca2+] of Fura2-loaded PMNs were detected. In the presence of extracellular Ca2+, the fluorescence of ethidium was not modified but the free intracellular [Ca2+] of PMNs increased after application of PVL in a concentration-dependent manner. The time lag observed before an increase in the ethidium fluorescence was longer than the time lag observed before a Fura2 fluorescence increase. Simultaneous recordings of the two probes fluorescence variations have shown the protective effect of Ca2+ and Zn2+ and the closing of the pore by 50 mM Ca2+ or 2 mM Zn2+. Moreover, the effect of Ca2+ could be reversed by the addition of EGTA. In the presence of 1 mM extracellular Ca2+ or 0.8 mM extracellular Zn2+, the pore induced by PVL had an ionic size allowing Ca2+, Mn2+, Zn2+ and Mg2+ fluxes. The addition of antibodies against either component of PVL inhibits the permeabilization provoked by the toxin even after it was initiated. It is concluded that leukocidin from S. aureus is a pore-forming toxin which, under physiological conditions ([Ca2+] = 1 to 1.5 mM), provokes the formation of an ion-sized pore inducing an increase in the free intracellular Ca2+ which may activate PMN functions.
Laboratoire de Toxinologie Bactérienne, Faculté de Médecine, Université Louis Pasteur, Strasbourg, France.
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Finck-Barbançon
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Duportail
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Meunier
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Colin
D002118:Calcium / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects; D002463:Cell Membrane Permeability / Q000187:drug effects; D004996:Ethidium; D016257:Fura-2; D006801:Humans; D007956:Leukocidins / Q000737:chemistry / Q000302:isolation & purification / Q000494:pharmacology*; D009504:Neutrophils / Q000187:drug effects* / Q000378:metabolism; D018272:Porins / Q000096:biosynthesis; D013211:Staphylococcus aureus / Q000737:chemistry*
D007956:Leukocidins; D018272:Porins; D004996:Ethidium; D002118:Calcium; D016257:Fura-2