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 c551, 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 c551, they underwent extensive cell death due to induction of apoptosis.
BuomminoE., MorelliF., MetaforaS., RossanoF., PerfetoB., BaroniA., TufanoM. A.1999; Porin from Pseudomonas aeruginosa induces apoptosis in an epithelial cell line derived from rat seminal vesicles. Infect Immun 67:4794–4800
CoburnJ., FrankD. W.1999; Macrophages and epithelial cells respond differently to the Pseudomonas aeruginosa type III secretion system. Infect Immun 67:3151–3154
DacheuxD., AttreeI., SchneiderC., ToussaintB.1999; Cell death of human polymorphonuclear neutrophils induced by a Pseudomonas aeruginosa cystic fibrosis isolate requires a functional type III secretion system. Infect Immun 67:6164–6167
DubyakG. R., El-MoatassimC.1993; Signal transduction via P2-purinergic receptors for extracellular ATP and other nucleotides. Am J Physiol 265:C577–C606
EllerbyH. M., MartinS. J., EllerbyL. M.7 other authors1997; Establishment of a cell-free system of neuronal apoptosis: comparison of premitochondrial, mitochondrial, and postmitochondrial phases. Neuroscience 17:6165–6178
FerrariD., ChiozziP., FalzoniS., HanauS., Di VirgilioF.1997; Purinergic modulation of interleukin-1β release from microglial cells stimulated with bacterial endotoxin. J Exp Med 185:579–582[CrossRef]
FerrariD., LosM., BaureM. K. A., VandenabeeleP., WeselborgS., Schulze-OsthoffK.1999; P2Z purinoreceptor ligation induces activation of caspases with distinct roles in apoptotic and necrotic alterations of cell death. FEBS Lett 447:71–75[CrossRef]
GoodhewC. F., BrownK. R., PettigrewG. W.1986; Haem staining in gels, a useful tool in the study of bacterial c-type cytochromes. Biochim Biphys Acta 852:288–294[CrossRef]
HershD., MonackD. M., SmithM. R., GhoriN., FalkowS., ZychlinskyA.1999; The Salmonella invasin SipB induces macrophage apoptosis by binding to caspase-1. Proc Natl Acad Sci USA 96:2396–2401[CrossRef]
HilbiH., MossJ. E., HershD.7 other authors1998; Shigella-induced apoptosis is dependent on caspase-1 which binds to IpaB. J Biol Chem 273:32895–32900[CrossRef]
LammasD. A., StoberC., HarveyC. J., KendrickN., PanchalinganS., KumararatneD. S.1997; ATP-induced killing of mycobacteria by human macrophages is mediated by purinergic P2Z (P2X7) receptors. Immunity 7:433–444[CrossRef]
VallisA. J., YahrT. L., BarbieriJ. T., FrankD. W.1999; Regulation of ExoS production and secretion by Pseudomonas aeruginosa in response to tissue culture conditions. Infect Immun 67:914–920
Van de KampM., SilverstriniM. C., BrunoriM., Van BeeumenJ., HaliF. C., CantersG. W.1990; Involvement of the hydrophobic patch of azurin in the electron-transfer reactions with cytochrome c551 and nitrite reductase. Eur J Biochem 194:109–118[CrossRef]
VijgenboomE., BuschJ. E., CantersG. W.1997; In vivo studies disprove an obligatory role of azurin in denitrification in Pseudomonas aeruginosa and show that azu expression is under control of RpoS and ANR. Microbiology 143:2853–2863[CrossRef]
ZouH., HenzelW. J., LiuX., LutschgA., WangX.1997; Apaf-1, a human protein homologous to C. elegans CED-4, participates in cytochrome c-dependent activation of caspase-3. Cell 90:405–413[CrossRef]
ZouH., LiY., LiuX., WangX.1999; An Apaf-1·cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9. J Biol Chem 274:11549–11556[CrossRef]
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