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Group B streptococcus GAPDH is released upon cell lysis, associates with bacterial surface, and induces apoptosis in murine macrophages.

ABSTRACT: Glyceraldehyde 3-phosphate dehydrogenases (GAPDH) are cytoplasmic glycolytic enzymes that, despite lacking identifiable secretion signals, have been detected at the surface of several prokaryotic and eukaryotic organisms where they exhibit non-glycolytic functions including adhesion to host components. Group B Streptococcus (GBS) is a human commensal bacterium that has the capacity to cause life-threatening meningitis and septicemia in newborns. Electron microscopy and fluorescence-activated cell sorter (FACS) analysis demonstrated the surface localization of GAPDH in GBS. By addressing the question of GAPDH export to the cell surface of GBS strain NEM316 and isogenic mutant derivatives of our collection, we found that impaired GAPDH presence in the surface and supernatant of GBS was associated with a lower level of bacterial lysis. We also found that following GBS lysis, GAPDH can associate to the surface of many living bacteria. Finally, we provide evidence for a novel function of the secreted GAPDH as an inducer of apoptosis of murine macrophages.

SUBMITTER: Oliveira L

PROVIDER: S-EPMC3264557 | biostudies-literature | 2012

REPOSITORIES: biostudies-literature

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Group B streptococcus GAPDH is released upon cell lysis, associates with bacterial surface, and induces apoptosis in murine macrophages.

Oliveira Liliana L Madureira Pedro P Andrade Elva Bonifácio EB Bouaboud Abdelouhab A Morello Eric E Ferreira Paula P Poyart Claire C Trieu-Cuot Patrick P Dramsi Shaynoor S

PloS one 20120123 1

Glyceraldehyde 3-phosphate dehydrogenases (GAPDH) are cytoplasmic glycolytic enzymes that, despite lacking identifiable secretion signals, have been detected at the surface of several prokaryotic and eukaryotic organisms where they exhibit non-glycolytic functions including adhesion to host components. Group B Streptococcus (GBS) is a human commensal bacterium that has the capacity to cause life-threatening meningitis and septicemia in newborns. Electron microscopy and fluorescence-activated cel ...[more]

PMID: 22291899

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Project description:UnlabelledAs key components of innate immune defense, macrophages are essential in controlling bacterial pathogens, including group A Streptococcus(GAS). Despite this, only a limited number of studies have analyzed the recovery of GAS from within human neutrophils and macrophages. Here, we determined the intracellular fate of GAS in human macrophages by using several quantitative approaches. In both U937 and primary human macrophages, the appearance over time of long GAS chains revealed that despite GAS-mediated cytotoxicity, replication occurred in viable, propidium iodide-negative macrophages. Whereas the major virulence factor M1 did not contribute to bacterial growth, a GAS mutant strain deficient in streptolysin O (SLO) was impaired for intracellular replication. SLO promoted bacterial escape from the GAS-containing vacuole (GCV) into the macrophage cytosol. Up to half of the cytosolic GAS colocalized with ubiquitin and p62, suggesting that the bacteria were targeted by the autophagy machinery. Despite this, live imaging of U937 macrophages revealed proficient replication of GAS after GCV rupture, indicating that escape from the GCV is important for growth of GAS in macrophages. Our results reveal that GAS can replicate within viable human macrophages, with SLO promoting GCV escape and cytosolic growth, despite the recruitment of autophagy receptors to bacteria.ImportanceClassically regarded as an extracellular pathogen, GAS can persist within human epithelial cells, as well as neutrophils and macrophages. Some studies suggest that GAS can modulate its intracellular vacuole to promote survival and perhaps replicate in macrophages. However, an in-depth single-cell analysis of the dynamics of survival and replication is lacking. We used macrophage-like cell lines and primary macrophages to measure the intracellular growth of GAS at both the population and single-cell levels. While CFU counts revealed no increase in overall bacterial growth, quantitative fluorescence microscopy, flow cytometry, and time-lapse imaging revealed bacterial replication in a proportion of infected macrophages. This study emphasizes the importance of single-cell analysis especially when studying the intracellular fate of a pathogen that is cytotoxic and displays heterogeneity in terms of intracellular killing and growth. To our knowledge, this study provides the first direct visualization of GAS replication inside human cells.

Project description:Group B Streptococcus (GBS) is the leading cause of neonatal pneumonia, septicemia, and meningitis. We have previously shown that in adult mice GBS glycolytic enzyme glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is an extracellular virulence factor that induces production of the immunosuppressive cytokine interleukin-10 (IL-10) by the host early upon bacterial infection. Here, we investigate whether immunity to neonatal GBS infection could be achieved through maternal vaccination against bacterial GAPDH. Female BALB/c mice were immunized with rGAPDH and the progeny was infected with a lethal inoculum of GBS strains. Neonatal mice born from mothers immunized with rGAPDH were protected against infection with GBS strains, including the ST-17 highly virulent clone. A similar protective effect was observed in newborns passively immunized with anti-rGAPDH IgG antibodies, or F(ab')(2) fragments, indicating that protection achieved with rGAPDH vaccination is independent of opsonophagocytic killing of bacteria. Protection against lethal GBS infection through rGAPDH maternal vaccination was due to neutralization of IL-10 production soon after infection. Consequently, IL-10 deficient (IL-10(-/-)) mice pups were as resistant to GBS infection as pups born from vaccinated mothers. We observed that protection was correlated with increased neutrophil trafficking to infected organs. Thus, anti-rGAPDH or anti-IL-10R treatment of mice pups before GBS infection resulted in increased neutrophil numbers and lower bacterial load in infected organs, as compared to newborn mice treated with the respective control antibodies. We showed that mothers immunized with rGAPDH produce neutralizing antibodies that are sufficient to decrease IL-10 production and induce neutrophil recruitment into infected tissues in newborn mice. These results uncover a novel mechanism for GBS virulence in a neonatal host that could be neutralized by vaccination or immunotherapy. As GBS GAPDH is a structurally conserved enzyme that is metabolically essential for bacterial growth in media containing glucose as the sole carbon source (i.e., the blood), this protein constitutes a powerful candidate for the development of a human vaccine against this pathogen.

Dataset Information

Group B streptococcus GAPDH is released upon cell lysis, associates with bacterial surface, and induces apoptosis in murine macrophages.

Publications

Group B streptococcus GAPDH is released upon cell lysis, associates with bacterial surface, and induces apoptosis in murine macrophages.

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