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alpha-Toxin is a mediator of Staphylococcus aureus-induced cell death and activates caspases via the intrinsic death pathway independently of death receptor signaling.

Bantel H, Sinha B, Domschke W, Peters G, Schulze-Osthoff K, Jänicke RU - J. Cell Biol. (2001)

Bottom Line: Furthermore, alpha-toxin-induced caspase activation in CD95-resistant Jurkat sublines lacking CD95, Fas-activated death domain, or caspase-8 but not in cells stably expressing the antiapoptotic protein Bcl-2.Together with our finding that alpha-toxin induces cytochrome c release in intact cells and, interestingly, also from isolated mitochondria in a Bcl-2-controlled manner, our results demonstrate that S. aureus alpha-toxin triggers caspase activation via the intrinsic death pathway independently of death receptors.Hence, our findings clearly define a signaling pathway used in S. aureus-induced cytotoxicity and may provide a molecular rationale for future therapeutic interventions in bacterial infections.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Cell Biology, University of Münster, 48149 Münster, Germany.

ABSTRACT
Infections with Staphylococcus aureus, a common inducer of septic and toxic shock, often result in tissue damage and death of various cell types. Although S. aureus was suggested to induce apoptosis, the underlying signal transduction pathways remained elusive. We show that caspase activation and DNA fragmentation were induced not only when Jurkat T cells were infected with intact bacteria, but also after treatment with supernatants of various S. aureus strains. We also demonstrate that S. aureus-induced cell death and caspase activation were mediated by alpha-toxin, a major cytotoxin of S. aureus, since both events were abrogated by two different anti-alpha-toxin antibodies and could not be induced with supernatants of an alpha-toxin-deficient S. aureus strain. Furthermore, alpha-toxin-induced caspase activation in CD95-resistant Jurkat sublines lacking CD95, Fas-activated death domain, or caspase-8 but not in cells stably expressing the antiapoptotic protein Bcl-2. Together with our finding that alpha-toxin induces cytochrome c release in intact cells and, interestingly, also from isolated mitochondria in a Bcl-2-controlled manner, our results demonstrate that S. aureus alpha-toxin triggers caspase activation via the intrinsic death pathway independently of death receptors. Hence, our findings clearly define a signaling pathway used in S. aureus-induced cytotoxicity and may provide a molecular rationale for future therapeutic interventions in bacterial infections.

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S. aureus–induced caspase-3 activation results in α-fodrin cleavage and DNA fragmentation. (A) Western blot analyses demonstrating caspase-3 processing (top) and α-fodrin cleavage (bottom) in Jurkat cells that were either left untreated (control) or incubated for 4 h with the indicated dilutions of Wood 46 supernatant (SN). (B) Agarose gel electrophoretic separation of DNA extracted from Jurkat cells that were either left untreated (control) or incubated for 6 h with the indicated dilutions of Wood 46 supernatant (SN). The positions of the DNA molecular size marker are indicated on the right.
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fig4: S. aureus–induced caspase-3 activation results in α-fodrin cleavage and DNA fragmentation. (A) Western blot analyses demonstrating caspase-3 processing (top) and α-fodrin cleavage (bottom) in Jurkat cells that were either left untreated (control) or incubated for 4 h with the indicated dilutions of Wood 46 supernatant (SN). (B) Agarose gel electrophoretic separation of DNA extracted from Jurkat cells that were either left untreated (control) or incubated for 6 h with the indicated dilutions of Wood 46 supernatant (SN). The positions of the DNA molecular size marker are indicated on the right.

Mentions: Western blot analysis confirmed these results, showing that treatment of Jurkat cells with supernatant of Wood 46 resulted in the rapid conversion of the inactive 32 kD caspase-3 precursor into the proteolytically active p17 subunit, indicating that caspase-3 was indeed activated during S. aureus–induced apoptosis (Fig. 4 A, top). Similar results were obtained when α-toxin was used as the death stimulus (unpublished data). In both cases, the appearance of the active p17 caspase-3 subunit was abrogated completely in the presence of the anti–α-toxin antibody (unpublished data), demonstrating that caspase-3 activation was induced by α-toxin. In agreement with the DEVDase assays (Figs. 3, A and C), caspase-3 activation peaked at lower Wood 46 concentrations (0.06% supernatant) and declined rapidly when higher concentrations were used (Fig. 4 A, top). Furthermore, caspase-3 activation correlated directly with cleavage of the death substrate α-fodrin into the 120-kD fragment (Fig. 4 A, bottom) and the appearance of the characteristic DNA ladder formation (Fig. 4 B), both of which could be inhibited completely by the anti–α-toxin antibody (unpublished data). Since it was shown recently that active caspase-3 is absolutely required for the induction of both these events (Jänicke et al., 1998a,b), our results clearly demonstrate that S. aureus–induced cell death and caspase activation are mediated by α-toxin.


alpha-Toxin is a mediator of Staphylococcus aureus-induced cell death and activates caspases via the intrinsic death pathway independently of death receptor signaling.

Bantel H, Sinha B, Domschke W, Peters G, Schulze-Osthoff K, Jänicke RU - J. Cell Biol. (2001)

S. aureus–induced caspase-3 activation results in α-fodrin cleavage and DNA fragmentation. (A) Western blot analyses demonstrating caspase-3 processing (top) and α-fodrin cleavage (bottom) in Jurkat cells that were either left untreated (control) or incubated for 4 h with the indicated dilutions of Wood 46 supernatant (SN). (B) Agarose gel electrophoretic separation of DNA extracted from Jurkat cells that were either left untreated (control) or incubated for 6 h with the indicated dilutions of Wood 46 supernatant (SN). The positions of the DNA molecular size marker are indicated on the right.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2198876&req=5

fig4: S. aureus–induced caspase-3 activation results in α-fodrin cleavage and DNA fragmentation. (A) Western blot analyses demonstrating caspase-3 processing (top) and α-fodrin cleavage (bottom) in Jurkat cells that were either left untreated (control) or incubated for 4 h with the indicated dilutions of Wood 46 supernatant (SN). (B) Agarose gel electrophoretic separation of DNA extracted from Jurkat cells that were either left untreated (control) or incubated for 6 h with the indicated dilutions of Wood 46 supernatant (SN). The positions of the DNA molecular size marker are indicated on the right.
Mentions: Western blot analysis confirmed these results, showing that treatment of Jurkat cells with supernatant of Wood 46 resulted in the rapid conversion of the inactive 32 kD caspase-3 precursor into the proteolytically active p17 subunit, indicating that caspase-3 was indeed activated during S. aureus–induced apoptosis (Fig. 4 A, top). Similar results were obtained when α-toxin was used as the death stimulus (unpublished data). In both cases, the appearance of the active p17 caspase-3 subunit was abrogated completely in the presence of the anti–α-toxin antibody (unpublished data), demonstrating that caspase-3 activation was induced by α-toxin. In agreement with the DEVDase assays (Figs. 3, A and C), caspase-3 activation peaked at lower Wood 46 concentrations (0.06% supernatant) and declined rapidly when higher concentrations were used (Fig. 4 A, top). Furthermore, caspase-3 activation correlated directly with cleavage of the death substrate α-fodrin into the 120-kD fragment (Fig. 4 A, bottom) and the appearance of the characteristic DNA ladder formation (Fig. 4 B), both of which could be inhibited completely by the anti–α-toxin antibody (unpublished data). Since it was shown recently that active caspase-3 is absolutely required for the induction of both these events (Jänicke et al., 1998a,b), our results clearly demonstrate that S. aureus–induced cell death and caspase activation are mediated by α-toxin.

Bottom Line: Furthermore, alpha-toxin-induced caspase activation in CD95-resistant Jurkat sublines lacking CD95, Fas-activated death domain, or caspase-8 but not in cells stably expressing the antiapoptotic protein Bcl-2.Together with our finding that alpha-toxin induces cytochrome c release in intact cells and, interestingly, also from isolated mitochondria in a Bcl-2-controlled manner, our results demonstrate that S. aureus alpha-toxin triggers caspase activation via the intrinsic death pathway independently of death receptors.Hence, our findings clearly define a signaling pathway used in S. aureus-induced cytotoxicity and may provide a molecular rationale for future therapeutic interventions in bacterial infections.

View Article: PubMed Central - PubMed

Affiliation: Department of Immunology and Cell Biology, University of Münster, 48149 Münster, Germany.

ABSTRACT
Infections with Staphylococcus aureus, a common inducer of septic and toxic shock, often result in tissue damage and death of various cell types. Although S. aureus was suggested to induce apoptosis, the underlying signal transduction pathways remained elusive. We show that caspase activation and DNA fragmentation were induced not only when Jurkat T cells were infected with intact bacteria, but also after treatment with supernatants of various S. aureus strains. We also demonstrate that S. aureus-induced cell death and caspase activation were mediated by alpha-toxin, a major cytotoxin of S. aureus, since both events were abrogated by two different anti-alpha-toxin antibodies and could not be induced with supernatants of an alpha-toxin-deficient S. aureus strain. Furthermore, alpha-toxin-induced caspase activation in CD95-resistant Jurkat sublines lacking CD95, Fas-activated death domain, or caspase-8 but not in cells stably expressing the antiapoptotic protein Bcl-2. Together with our finding that alpha-toxin induces cytochrome c release in intact cells and, interestingly, also from isolated mitochondria in a Bcl-2-controlled manner, our results demonstrate that S. aureus alpha-toxin triggers caspase activation via the intrinsic death pathway independently of death receptors. Hence, our findings clearly define a signaling pathway used in S. aureus-induced cytotoxicity and may provide a molecular rationale for future therapeutic interventions in bacterial infections.

Show MeSH
Related in: MedlinePlus