Effect of staphylococcal alpha-toxin on intracellular Ca2+ in polymorphonuclear leukocytes.
Staphylococcal alpha-toxin, a channel-forming protein, stimulates leukotriene B4 formation in rabbit polymorphonuclear leukocytes (PMN) (N. Suttorp, W. Seeger, J. Zucker-Reimann, L. Roka, and S. Bhakdi, Infect. Immun. 55:104-110, 1987). The concept was advanced that transmembrane toxin pores act as Ca2+ gates allowing passive Ca2+ influx into the cell, thus initiating stimulus response coupling. A critical step in this hypothesis is the demonstration of an increase in the cytosolic free Ca2+ concentration [( Ca2+]i). [Ca2+]i and membrane-associated Ca2+ were therefore monitored in quin-2- or chlorotetracycline-loaded PMN exposed to alpha-toxin. The effects of the Ca2+ ionophore ionomycin and the chemotactic tripeptide formylmethionyl-leucylphenylalanine (fMLP) were studied in parallel. All stimuli increased [Ca2+]i in dose- and time-dependent manner. In the presence of an EDTA excess there was a decrease of [Ca2+]i due to an efflux of Ca2+ in alpha-toxin- and ionomycin-treated cells, while addition of fMLP still induced an increase of [Ca2+]i. In the presence of verapamil, a Ca2+ channel blocker, [Ca2+]i was reduced after stimulation with fMLP but not with alpha-toxin or ionomycin. Addition of fMLP and ionomycin but not of alpha-toxin to PMN resulted in a rapid and substantial mobilization of membrane-associated Ca2+. The collective data demonstrate that exposure of PMN to staphylococcal alpha-toxin results in an increase in [Ca2+]i which is due to an influx of extracellular Ca2+ and not to a mobilization of intracellularly stored Ca2+. The concept of initiating stimulus response coupling by Ca2+ influx through transmembrane pores may be generally applicable to other channel-forming cytolysins.
D000818:Animals; D001427:Bacterial Toxins / Q000494:pharmacology*; D001826:Body Fluids / Q000378:metabolism*; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000382:microbiology; D004492:Edetic Acid / Q000494:pharmacology; D004987:Ethers / Q000494:pharmacology; D006460:Hemolysin Proteins; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism* / Q000382:microbiology; D015759:Ionomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism* / Q000382:microbiology; D011817:Rabbits; D013211:Staphylococcus aureus / Q000472:pathogenicity; D014700:Verapamil / Q000378:metabolism
D001427:Bacterial Toxins; D004987:Ethers; D006460:Hemolysin Proteins; C021091:staphylococcal alpha-toxin; D015759:Ionomycin; D009240:N-Formylmethionine Leucyl-Phenylalanine; D004492:Edetic Acid; D014700:Verapamil