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Neutrophils are resistant to Yersinia YopJ/P-induced apoptosis and are protected from ROS-mediated cell death by the type III secretion system.

Spinner JL, Seo KS, O'Loughlin JL, Cundiff JA, Minnich SA, Bohach GA, Kobayashi SD - PLoS ONE (2010)

Bottom Line: In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death.PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner.Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Molecular Biology and Biochemistry, University of Idaho, Moscow, Idaho, United States of America.

ABSTRACT

Background: The human innate immune system relies on the coordinated activity of macrophages and polymorphonuclear leukocytes (neutrophils or PMNs) for defense against bacterial pathogens. Yersinia spp. subvert the innate immune response to cause disease in humans. In particular, the Yersinia outer protein YopJ (Y. pestis and Y. pseudotuberculosis) and YopP (Y. enterocolitica) rapidly induce apoptosis in murine macrophages and dendritic cells. However, the effects of Yersinia Yop J/P on neutrophil fate are not clearly defined.

Methodology/principal findings: In this study, we utilized wild-type and mutant strains of Yersinia to test the contribution of YopJ and YopP on induction of apoptosis in human monocyte-derived macrophages (HMDM) and neutrophils. Whereas YopJ and YopP similarly induced apoptosis in HMDMs, interaction of human neutrophils with virulence plasmid-containing Yersinia did not result in PMN caspase activation, release of LDH, or loss of membrane integrity greater than PMN controls. In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death. PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner. Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

Conclusions: Our findings showed that Yersinia YopJ and/or YopP did not induce pronounced apoptosis in human neutrophils. Furthermore, robust PMN ROS production in response to virulence plasmid-deficient Yersinia was associated with increased PMN cell death, suggesting that Yersinia inhibition of PMN ROS production plays a role in evasion of the human innate immune response in part by limiting PMN apoptosis.

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Induction of caspase activity is YopJ/YopP dependent but delayed in human MDMs compared to J774A.1 cells.HMDMs and J774A.1 cells were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units (RFLU). Results are expressed as the mean ± SEM of three experiments. *, represents difference from HMDM and J774A.1 controls (P<0.05).
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pone-0009279-g002: Induction of caspase activity is YopJ/YopP dependent but delayed in human MDMs compared to J774A.1 cells.HMDMs and J774A.1 cells were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units (RFLU). Results are expressed as the mean ± SEM of three experiments. *, represents difference from HMDM and J774A.1 controls (P<0.05).

Mentions: To assess the association of YopJ and YopP with caspase activation in macrophages, we measured caspase-3, -8, -9, and -2 activity following 3 h (data not shown) and 6 h incubation with Yersinia. All caspases tested were rapidly activated in J774A.1 macrophages after incubation with KIM5, Y. enterocolitica, or KIM5 YopJ-YopP but not KIM6 or KIM5ΔyopJ (Fig. 2). HMDMs incubated with Yersinia did not result in activation of any caspase tested until ∼6 h (Fig. 2). At 6 h, all caspases tested were activated in HMDMs by YopJ/YopP expressing strains but not by KIM6 or KIM5ΔyopJ (Fig. 2). Delayed caspase activation by YopJ/YopP in HMDMs relative to J774A.1 cells corresponds with the observed delays in HMDM lysis and EthD-1 staining (Fig. 1).


Neutrophils are resistant to Yersinia YopJ/P-induced apoptosis and are protected from ROS-mediated cell death by the type III secretion system.

Spinner JL, Seo KS, O'Loughlin JL, Cundiff JA, Minnich SA, Bohach GA, Kobayashi SD - PLoS ONE (2010)

Induction of caspase activity is YopJ/YopP dependent but delayed in human MDMs compared to J774A.1 cells.HMDMs and J774A.1 cells were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units (RFLU). Results are expressed as the mean ± SEM of three experiments. *, represents difference from HMDM and J774A.1 controls (P<0.05).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2823771&req=5

pone-0009279-g002: Induction of caspase activity is YopJ/YopP dependent but delayed in human MDMs compared to J774A.1 cells.HMDMs and J774A.1 cells were incubated alone or with Y. pestis KIM5, KIM5 YopJ-YopP, KIM5ΔyopJ, KIM6 and Y. enterocolitica 8081v grown at 37°C. Caspase-3, -8, -9, and -2 activity was measured after 6 h of incubation as described in Materials and Methods and is expressed in relative fluorescence units (RFLU). Results are expressed as the mean ± SEM of three experiments. *, represents difference from HMDM and J774A.1 controls (P<0.05).
Mentions: To assess the association of YopJ and YopP with caspase activation in macrophages, we measured caspase-3, -8, -9, and -2 activity following 3 h (data not shown) and 6 h incubation with Yersinia. All caspases tested were rapidly activated in J774A.1 macrophages after incubation with KIM5, Y. enterocolitica, or KIM5 YopJ-YopP but not KIM6 or KIM5ΔyopJ (Fig. 2). HMDMs incubated with Yersinia did not result in activation of any caspase tested until ∼6 h (Fig. 2). At 6 h, all caspases tested were activated in HMDMs by YopJ/YopP expressing strains but not by KIM6 or KIM5ΔyopJ (Fig. 2). Delayed caspase activation by YopJ/YopP in HMDMs relative to J774A.1 cells corresponds with the observed delays in HMDM lysis and EthD-1 staining (Fig. 1).

Bottom Line: In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death.PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner.Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

View Article: PubMed Central - PubMed

Affiliation: Department of Microbiology, Molecular Biology and Biochemistry, University of Idaho, Moscow, Idaho, United States of America.

ABSTRACT

Background: The human innate immune system relies on the coordinated activity of macrophages and polymorphonuclear leukocytes (neutrophils or PMNs) for defense against bacterial pathogens. Yersinia spp. subvert the innate immune response to cause disease in humans. In particular, the Yersinia outer protein YopJ (Y. pestis and Y. pseudotuberculosis) and YopP (Y. enterocolitica) rapidly induce apoptosis in murine macrophages and dendritic cells. However, the effects of Yersinia Yop J/P on neutrophil fate are not clearly defined.

Methodology/principal findings: In this study, we utilized wild-type and mutant strains of Yersinia to test the contribution of YopJ and YopP on induction of apoptosis in human monocyte-derived macrophages (HMDM) and neutrophils. Whereas YopJ and YopP similarly induced apoptosis in HMDMs, interaction of human neutrophils with virulence plasmid-containing Yersinia did not result in PMN caspase activation, release of LDH, or loss of membrane integrity greater than PMN controls. In contrast, interaction of human PMNs with the virulence plasmid-deficient Y. pestis strain KIM6 resulted in increased surface exposure of phosphatidylserine (PS) and cell death. PMN reactive oxygen species (ROS) production was inhibited in a virulence plasmid-dependent but YopJ/YopP-independent manner. Following phagocytic interaction with Y. pestis strain KIM6, inhibition of PMN ROS production with diphenyleneiodonium chloride resulted in a reduction of PMN cell death similar to that induced by the virulence plasmid-containing strain Y. pestis KIM5.

Conclusions: Our findings showed that Yersinia YopJ and/or YopP did not induce pronounced apoptosis in human neutrophils. Furthermore, robust PMN ROS production in response to virulence plasmid-deficient Yersinia was associated with increased PMN cell death, suggesting that Yersinia inhibition of PMN ROS production plays a role in evasion of the human innate immune response in part by limiting PMN apoptosis.

Show MeSH
Related in: MedlinePlus