AEROLYSIN INDUCES G-PROTEIN ACTIVATION AND CA2+ RELEASE FROM INTRACELLULAR STORES IN HUMAN GRANULOCYTES

Aerolysin is a pore-forming toxin that plays a key role in the pathoge nesis of Aeromonas hydrophila infections. In this study, we have analy zed the effect of aerolysin on human granulocytes (HL-60 cells). Proae rolysin could bind to these cells, was processed into active aerolysin , and led to membrane depolarization, indicating that granulocytes are potential targets for this toxin, Fura-a measurements were used to an alyze the effect of aerolysin lysin on cytosolic [Ca2+] homeostasis. A s expected for a pore-forming toxin, aerolysin addition led to Ca2+ in flux across the plasma membrane. In addition, the toxin triggered Ca2 release from agonist and thapsigargin-sensitive intracellular Ca2+ st ores. This Ca2+ release was independent of the aerolysin-induced Ca2influx and occurred in two kinetically distinct phases: an initial rap id and transient phase and a second, more sustained, phase. The first, but not the second phase was sensitive to pertussis toxin, Activation of pertussis toxin-sensitive G-proteins appeared to be a consequence of pore formation, rather than receptor activation through aerolysin-b inding, as ik (i) was not observed with a binding competent, insertion -incompetent aerolysin mutant, (ii) had a marked lag time, and (iii) w as also observed in response to other bacterial pore-forming toxins (s taphylococcal alpha-toxin, streptolysin O) which are thought to bind t o different receptors, G-protein activation through pore-forming toxin s stimulated cellular functions, as evidenced by pertussis toxin-sensi tive chemotaxis. Our results demonstrate that granulocytes are potenti al target cells for aerolysin and that in these cells, Ca2+ signaling in response to a pore-forming toxin involves G-protein-dependent cell activation and Ca2+ release from intracellular stores.