Staphylococcus aureus alpha-toxin inhibits CD8+ T cell-mediated killing of cancer cells in cutaneous T-cell lymphoma.
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ABSTRACT:
SUBMITTER: Blumel E
PROVIDER: S-EPMC7185203 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Staphylococcus aureus is a major cause of community-acquired and nosocomial infections including the life-threatening conditions endocarditis, necrotizing pneumonia, necrotizing fasciitis, and septicemia. Toll-like receptor (TLR)-2, a membrane-bound microbial sensor, detects staphylococcal components, but macrophages lacking TLR2 or both TLR2 and TLR4 remain S. aureus responsive, suggesting that an alternative microbial recognition receptor might be involved. The cytoplasmic sensor nucleotide-binding oligomerization domain containing (NOD) 2/caspase recruitment domain (CARD) 15 detects muramyl dipeptide from bacterial peptidoglycans and mediates cytokine responses to S. aureus in vitro, but the physiological significance of these observations is not well defined. Here we show that NOD2-deficient mice exhibit a delayed but ultimately exacerbated ulcerative response and impaired bacterial clearance after s.c. infection with S. aureus. NOD2-dependent recognition of S. aureus and muramyl dipeptide is facilitated by alpha-toxin (alpha-hemolysin), a pore-forming toxin and virulence factor of the pathogen. The action of NOD2 is dependent on IL-1beta-amplified production of IL-6, which promotes rapid bacterial killing by neutrophils. These results significantly broaden the physiological importance of NOD2 in innate immunity from the recognition of bacteria that primarily enter the cytoplasm to the detection of bacteria that typically reside extracellularly and demonstrate that this microbial sensor contributes to the discrimination between commensal bacteria and bacterial pathogens that elaborate pore-forming toxins.
| S-EPMC2722361 | biostudies-literature
Project description:Staphylococcus aureus is implicated in disease progression in cutaneous T-cell lymphoma (CTCL). Here, we demonstrate that malignant T cell lines derived from CTCL patients as well as primary malignant CD4+ T cells from Sézary syndrome patients are considerably more resistant to alpha-toxin-induced cell death than their non-malignant counterparts. Thus, in a subset of Sézary syndrome patients the ratio between malignant and non-malignant CD4+ T cells increases significantly following exposure to alpha-toxin. Whereas toxin-induced cell death is ADAM10 dependent in healthy CD4+ T cells, resistance to alpha-toxin in malignant T cells involves both downregulation of ADAM10 as well as other resistance mechanisms. In conclusion, we provide first evidence that Staphylococcus aureus derived alpha-toxin can tilt the balance between malignant and non-malignant CD4+ T cells in CTCL patients. Consequently, alpha-toxin may promote disease progression through positive selection of malignant CD4+ T cells, identifying alpha-toxin as a putative drug target in CTCL.
| S-EPMC6791457 | biostudies-literature
Project description:Resistance and tolerance are two defense strategies employed by the host against microbial threats. Autophagy-mediated degradation of bacteria has been extensively described as a major resistance mechanism. Here we find that the dominant function of autophagy proteins during infections with the epidemic community-associated methicillin-resistant Staphylococcus aureus USA300 is to mediate tolerance rather than resistance. Atg16L1 hypomorphic mice (Atg16L1(HM)), which have reduced autophagy, were highly susceptible to lethality in both sepsis and pneumonia models of USA300 infection. Autophagy confers protection by limiting the damage caused by ?-toxin, particularly to endothelial cells. Remarkably, Atg16L1(HM) mice display enhanced survival rather than susceptibility upon infection with ?-toxin-deficient S. aureus. These results identify an essential role for autophagy in tolerance to Staphylococcal disease and highlight how a single virulence factor encoded by a pathogen can determine whether a given host factor promotes tolerance or resistance.
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Project description:Staphylococcus aureus infection is one of the leading causes of morbidity in hospitalized patients in the United States, an effect compounded by increasing antibiotic resistance. The secreted agent hemolysin alpha toxin (Hla) requires the receptor A Disintegrin And Metalloproteinase domain-containing protein 10 (ADAM10) to mediate its toxic effects. We hypothesized that these effects are in part regulated by Notch signaling, for which ADAM10 activation is essential. Notch proteins function in developmental and pathological angiogenesis via the modulation of key pathways in endothelial and perivascular cells. Thus, we hypothesized that Hla would activate Notch in vascular cells. Human umbilical vein endothelial cells were treated with recombinant Hla (rHla), Hla-H35L (genetically inactivated Hla), or Hank's solution (HBSS), and probed by different methods. Luciferase assays showed that Hla (0.01 µg/mL) increased Notch activation by 1.75?±?0.5-fold as compared to HBSS controls (p?<?0.05), whereas Hla-H35L had no effect. Immunocytochemistry and Western blotting confirmed these findings and revealed that ADAM10 and ?-secretase are required for Notch activation after inhibitor and siRNA assays. Retinal EC in mice engineered to express yellow fluorescent protein (YFP) upon Notch activation demonstrated significantly greater YFP intensity after Hla injection than controls. Aortic rings from Notch reporter mice embedded in matrix and incubated with rHla or Hla-H35L demonstrate increased Notch activation occurs at tip cells during sprouting. These mice also had higher skin YFP intensity and area of expression after subcutaneous inoculation of S. aureus expressing Hla than a strain lacking Hla in both EC and pericytes assessed by microscopy. Human liver displayed strikingly higher Notch expression in EC and pericytes during S. aureus infection by immunohistochemistry than tissues from uninfected patients. In sum, our results demonstrate that the S. aureus toxin Hla can potently activate Notch in vascular cells, an effect which may contribute to the pathobiology of infection with this microorganism.
| S-EPMC6360126 | biostudies-literature
Project description:Biofilms complicate treatment of Staphylococcus aureus (SA) wound infections. Previously, we determined alpha-toxin (AT)-promoted SA biofilm formation on mucosal tissue. Therefore, we evaluated SA wound isolates for AT production and biofilm formation on epithelium and assessed the role of AT in biofilm formation. Thirty-eight wound isolates were molecularly typed by pulsed-field gel electrophoresis (PFGE), multilocus sequence typing (ST), and spa typing. We measured biofilm formation of these SA isolates in vitro and ex vivo and quantified ex vivo AT production. We also investigated the effect of an anti-AT monoclonal antibody (MEDI4893*) on ex vivo biofilm formation by methicillin-resistant SA (USA 300 LAC) and tested whether purified AT rescued the biofilm defect of hla mutant SA strains. The predominant PFGE/ST combinations were USA100/ST5 (50%) and USA300/ST8 (33%) for methicillin-resistant SA (MRSA, n = 18), and USA200/ST30 (20%) for methicillin-susceptible SA (MSSA, n = 20). Ex vivo AT production correlated significantly with ex vivo SA wound isolate biofilm formation. Anti-alpha-toxin monoclonal antibody (MEDI4893*) prevented ex vivo biofilm formation by MRSA USA300 strain LAC. Wild-type AT rescued the ex vivo biofilm defect of non-AT producing SA strains. These findings provide evidence that AT plays a role in SA biofilm formation on epithelial surfaces and suggest that neutralization of AT may be useful in preventing and treating SA infections.
| S-EPMC5923323 | biostudies-literature
Project description:Staphylococcus aureus (S. aureus) is well known to express a plethora of toxins of which the pore-forming hemolysin A (α-toxin) is the best-studied cytolysin. Pore-forming toxins (PFT) permeabilize host membranes during infection thereby causing concentration-dependent effects in host cell membranes ranging from disordered ion fluxes to cytolysis. Host cells possess defense mechanisms against PFT attack, resulting in endocytosis of the breached membrane area and delivery of repair vesicles to the insulted plasma membrane as well as a concurrent release of membrane repair enzymes. Since PFTs from several pathogens have been shown to recruit membrane repair components, we here investigated whether staphylococcal α-toxin is able to induce these mechanisms in endothelial cells. We show that S. aureus α-toxin induced increase in cytosolic Ca2+ in endothelial cells, which was accompanied by p38 MAPK phosphorylation. Toxin challenge led to increased endocytosis of an extracellular fluid phase marker as well as increased externalization of LAMP1-positive membranes suggesting that peripheral lysosomes are recruited to the insulted plasma membrane. We further observed that thereby the lysosomal protein acid sphingomyelinase (ASM) was released into the cell culture medium. Thus, our results show that staphylococcal α-toxin triggers mechanisms in endothelial cells, which have been implicated in membrane repair after damage of other cell types by different toxins.
| S-EPMC8358437 | biostudies-literature
Project description:Staphylococcus aureus is a leading cause of human bacterial infection, causing a wide spectrum of disease ranging from skin and soft tissue infections to life-threatening pneumonia and sepsis. S. aureus toxins play an essential role in disease pathogenesis, contributing to both immunomodulation and host tissue injury. Prominent among these toxins are the membrane-active pore-forming cytolysin alpha-toxin (Hla) and the amphipathic ?-helical phenol-soluble modulin (PSM) peptides. As deletion of either the hla or psm locus leads to a phenotypically similar virulence defect in skin and soft tissue infection, we sought to determine the relative contribution of each locus to disease pathogenesis. Here we show that production of Hla can be modulated by PSM expression. An S. aureus mutant lacking PSM expression exhibits a transcriptional delay in hla mRNA production and therefore fails to secrete normal levels of Hla at early phases of growth. This leads to attenuation of virulence in vitro and in murine skin and lung models of infection, correlating with reduced recovery of Hla from host tissues. Production of Hla and restoration of staphylococcal virulence can be achieved in the psm mutant by plasmid-driven overexpression of hla. Our study suggests the coordinated action of Hla and PSMs in host tissue during early pathogenesis, confirming a major role for Hla in epithelial injury during S. aureus infection. These findings highlight the possibility that therapeutics targeting PSM production may simultaneously prevent Hla-mediated tissue injury.
| S-EPMC4136214 | biostudies-literature
Project description:The macrophage response to planktonic Staphylococcus aureus involves the induction of proinflammatory microbicidal activity. However, S. aureus biofilms can interfere with these responses in part by polarizing macrophages toward an anti-inflammatory profibrotic phenotype. Here we demonstrate that conditioned medium from mature S. aureus biofilms inhibited macrophage phagocytosis and induced cytotoxicity, suggesting the involvement of a secreted factor(s). Iterative testing found the active factor(s) to be proteinaceous and partially agr-dependent. Quantitative mass spectrometry identified alpha-toxin (Hla) and leukocidin AB (LukAB) as critical molecules secreted by S. aureus biofilms that inhibit murine macrophage phagocytosis and promote cytotoxicity. A role for Hla and LukAB was confirmed by using hla and lukAB mutants, and synergy between the two toxins was demonstrated with a lukAB hla double mutant and verified by complementation. Independent confirmation of the effects of Hla and LukAB on macrophage dysfunction was demonstrated by using an isogenic strain in which Hla was constitutively expressed, an Hla antibody to block toxin activity, and purified LukAB peptide. The importance of Hla and LukAB during S. aureus biofilm formation in vivo was assessed by using a murine orthopedic implant biofilm infection model in which the lukAB hla double mutant displayed significantly lower bacterial burdens and more macrophage infiltrates than each single mutant. Collectively, these findings reveal a critical synergistic role for Hla and LukAB in promoting macrophage dysfunction and facilitating S. aureus biofilm development in vivo. Staphylococcus aureus has a propensity to form multicellular communities known as biofilms. While growing in a biofilm, S. aureus displays increased tolerance to nutrient deprivation, antibiotic insult, and even host immune challenge. Previous studies have shown that S. aureus biofilms thwart host immunity in part by preventing macrophage phagocytosis. It remained unclear whether this was influenced solely by the considerable size of biofilms or whether molecules were also actively secreted to circumvent macrophage-mediated phagocytosis. This is the first report to demonstrate that S. aureus biofilms inhibit macrophage phagocytosis and induce macrophage death through the combined action of leukocidin AB and alpha-toxin. Loss of leukocidin AB and alpha-toxin expression resulted in enhanced S. aureus biofilm clearance in a mouse model of orthopedic implant infection, suggesting that these toxins could be targeted therapeutically to facilitate biofilm clearance in humans.
| S-EPMC4550693 | biostudies-literature
Project description:Staphylococcus aureus remains a leading cause of human infection. These infections frequently recur when the skin is a primary site of infection, especially in infants and children. In contrast, invasive staphylococcal disease is less commonly associated with reinfection, suggesting that tissue-specific mechanisms govern the development of immunity. Knowledge of how S. aureus manipulates protective immunity has been hampered by a lack of antigen-specific models to interrogate the T cell response. Using a chicken egg OVA-expressing S. aureus strain to analyze OVA-specific T cell responses, we demonstrated that primary skin infection was associated with impaired development of T cell memory. Conversely, invasive infection induced antigen-specific memory and protected against reinfection. This defect in adaptive immunity following skin infection was associated with a loss of DCs, attributable to S. aureus ?-toxin (Hla) expression. Gene- and immunization-based approaches to protect against Hla during skin infection restored the T cell response. Within the human population, exposure to ?-toxin through skin infection may modulate the establishment of T cell-mediated immunity, adversely affecting long-term protection. These studies prompt consideration that vaccination targeting S. aureus may be most effective if delivered prior to initial contact with the organism.
| S-EPMC7269593 | biostudies-literature
Project description:Staphylococcus aureus alpha-hemolysin (Hla) assembles into heptameric pores on the host cell membrane, causing lysis, apoptosis, and junction disruption. Herein, we present the design of a newly engineered S. aureus alpha-toxin, HlaPSGS, which lacks the predicted membrane-spanning stem domain. This protein is able to form heptamers in aqueous solution in the absence of lipophilic substrata, and its structure, obtained by transmission electron microscopy and single-particle reconstruction analysis, resembles the cap of the wild-type cytolytic Hla pore. HlaPSGS was found to be impaired in binding to host cells and to its receptor ADAM10 and to lack hemolytic and cytotoxic activity. Immunological studies using human sera as well as sera from mice convalescent from S. aureus infection suggested that the heptameric conformation of HlaPSGS mimics epitopes exposed by the cytolytic Hla pore during infection. Finally, immunization with this newly engineered Hla generated high protective immunity against staphylococcal infection in mice. Overall, this study provides unprecedented data on the natural immune response against Hla and suggests that the heptameric HlaPSGS is a highly valuable vaccine candidate against S. aureus.
| S-EPMC4895005 | biostudies-literature