Secreted products of a nonmucoid Pseudomonas aeruginosa strain induce two modes of macrophage killing: external-ATP-dependent, P2Z-receptor-mediated necrosis and ATP-independent, caspase-mediated apoptosis.
A nonmucoid clinical isolate of Pseudomonas aeruginosa, strain 808, elaborated ATP-dependent and ATP-independent types of cytotoxic factors in the growth medium. These cytotoxic factors, active against macrophages, were secreted during the exponential phase of growth in a complex medium. Commensurate with the appearance of the cytotoxic activities in the cell-free growth medium, several ATP-utilizing enzymic activities, such as adenylate kinase, nucleoside diphosphate kinase and 5'-nucleotidase (ATPase and/or phosphatase), were detected in the medium. These ATP-utilizing enzymes are believed to convert external ATP, presumably effluxed from macrophages, to various adenine nucleotides, which then activate purinergic receptors such as P2Z, leading to enhanced macrophage cell death. Pretreatment of macrophages with periodate-oxidized ATP (oATP), which is an irreversible inhibitor of P2Z receptor activation, prevented subsequent ATP-induced macrophage cell death. A second type of cytotoxic factor(s) operated in an ATP-independent manner such that it triggered activation of apoptotic processes in macrophages, leading to proteolytic conversion of procaspase-3 to active caspase-3. This cytotoxic factor(s) did not appear to act on procaspase-3 present in macrophage cytosolic extracts. Intact macrophages, when exposed to the cytotoxic factor(s) for 6-16 h, underwent apoptosis and demonstrated the presence of active caspase-3 in their cytosolic extracts. Interestingly, two redox proteins, azurin and cytochrome c(551), were detected in the cytotoxic preparation. When cell-line-derived or peritoneal macrophages or mast cells were incubated overnight with Q-Sepharose column flow-through fraction or with a mixture of azurin and cytochrome c(551), they underwent extensive cell death due to induction of apoptosis.
Zaborina
Olga
O
Dept of Microbiology & Immunology1 and Research Resource Center2, University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612, USA.
Dhiman
Neelam
N
Dept of Microbiology & Immunology1 and Research Resource Center2, University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612, USA.
Ling Chen
Mei
M
Dept of Microbiology & Immunology1 and Research Resource Center2, University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612, USA.
Kostal
Jan
J
Dept of Microbiology & Immunology1 and Research Resource Center2, University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612, USA.
Holder
Ian Alan
IA
Dept of Microbiology, Shriners Burns Hospital, 3229 Burnet Avenue, Cincinnati, OH 45229, USA3.
Chakrabarty
Ananda M
AM
Dept of Microbiology & Immunology1 and Research Resource Center2, University of Illinois College of Medicine, 835 South Wolcott Avenue, Chicago, IL 60612, USA.
D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D001400:Azurin / Q000494:pharmacology; D053148:Caspase 3; D020169:Caspases / Q000378:metabolism*; D002460:Cell Line; D002478:Cells, Cultured; D003578:Cytochrome-c Peroxidase / Q000494:pharmacology; D004798:Enzymes / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000502:physiology*; D017737:Macrophages, Peritoneal / Q000502:physiology; D008407:Mast Cells / Q000502:physiology; D051379:Mice; D009336:Necrosis; D011552:Pseudomonas Infections / Q000382:microbiology; D011550:Pseudomonas aeruginosa / Q000472:pathogenicity* / Q000502:physiology; D018048:Receptors, Purinergic P2 / Q000378:metabolism*; D058486:Receptors, Purinergic P2X7
D004798:Enzymes; C546468:P2RX7 protein, human; C546469:P2rx7 protein, mouse; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D001400:Azurin; D000255:Adenosine Triphosphate; D003578:Cytochrome-c Peroxidase; C505407:CASP3 protein, human; C505409:Casp3 protein, mouse; D053148:Caspase 3; D020169:Caspases