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DNase Sda1 allows invasive M1T1 Group A Streptococcus to prevent TLR9-dependent recognition.

Uchiyama S, Andreoni F, Schuepbach RA, Nizet V, Zinkernagel AS - PLoS Pathog. (2012)

Bottom Line: Similarly, in a murine necrotizing fasciitis model, IFN-α and TNF-α levels were significantly decreased in wild type mice infected with GAS expressing Sda1, whereas no such Sda1-dependent effect was seen in a TLR9-deficient background.Thus GAS Sda1 suppressed both the TLR9-mediated innate immune response and macrophage bactericidal activity.Our results demonstrate a novel mechanism of bacterial innate immune evasion based on autodegradation of CpG-rich DNA by a bacterial DNase.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.

ABSTRACT
Group A Streptococcus (GAS) has developed a broad arsenal of virulence factors that serve to circumvent host defense mechanisms. The virulence factor DNase Sda1 of the hyperinvasive M1T1 GAS clone degrades DNA-based neutrophil extracellular traps allowing GAS to escape extracellular killing. TLR9 is activated by unmethylated CpG-rich bacterial DNA and enhances innate immune resistance. We hypothesized that Sda1 degradation of bacterial DNA could alter TLR9-mediated recognition of GAS by host innate immune cells. We tested this hypothesis using a dual approach: loss and gain of function of DNase in isogenic GAS strains and presence and absence of TLR9 in the host. Either DNA degradation by Sda1 or host deficiency of TLR9 prevented GAS induced IFN-α and TNF-α secretion from murine macrophages and contributed to bacterial survival. Similarly, in a murine necrotizing fasciitis model, IFN-α and TNF-α levels were significantly decreased in wild type mice infected with GAS expressing Sda1, whereas no such Sda1-dependent effect was seen in a TLR9-deficient background. Thus GAS Sda1 suppressed both the TLR9-mediated innate immune response and macrophage bactericidal activity. Our results demonstrate a novel mechanism of bacterial innate immune evasion based on autodegradation of CpG-rich DNA by a bacterial DNase.

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Stimulation of murine macrophages by human and GAS DNA.(A–C) Bone marrow-derived mice macrophages (BMDMs) were stimulated with GAS and human genomic DNA (both 5 µg/ml) and secretion of IFN1, IFN-α and TNF-α in the supernatants measured. (D) Dose-dependency of GAS DNA-mediated stimulation (12 hours) of IFN-α and TNF-α secretion. Data were pooled from 3 experiments done in triplicates and presented as mean ± SEM. * P<0.05 ** P<0.01.
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ppat-1002736-g001: Stimulation of murine macrophages by human and GAS DNA.(A–C) Bone marrow-derived mice macrophages (BMDMs) were stimulated with GAS and human genomic DNA (both 5 µg/ml) and secretion of IFN1, IFN-α and TNF-α in the supernatants measured. (D) Dose-dependency of GAS DNA-mediated stimulation (12 hours) of IFN-α and TNF-α secretion. Data were pooled from 3 experiments done in triplicates and presented as mean ± SEM. * P<0.05 ** P<0.01.

Mentions: DNA was purified from GAS strain 5448, representative of the globally disseminated hyperinvasive M1T1 clone, and incubated with murine bone marrow-derived macrophages (BMDMs). Cytokine release into the medium was used as readout for BMDM activation. GAS DNA induced time-dependent release of interferon type 1 (IFN-1), and specifically interferon-α (IFN-α), from the macrophages (Fig. 1A, B), peaking at 12 h of exposure. GAS DNA also induced BMDM TNF-α secretion, with maximal levels already detected after 6 h of incubation and remaining elevated for at least 24 h (Fig. 1C). Induction of IFN-α and TNF-α release by GAS DNA was also dose-dependent (Fig. 1D). In contrast, human DNA did not induce IFN-1 or TNF-α secretion from BMDMs (Fig. 1A–C). We did not detect specific induction of the cytokines IL-6, IL-1β, IL-10 or MIP-2 from BMDMs exposed to GAS DNA (data not shown).


DNase Sda1 allows invasive M1T1 Group A Streptococcus to prevent TLR9-dependent recognition.

Uchiyama S, Andreoni F, Schuepbach RA, Nizet V, Zinkernagel AS - PLoS Pathog. (2012)

Stimulation of murine macrophages by human and GAS DNA.(A–C) Bone marrow-derived mice macrophages (BMDMs) were stimulated with GAS and human genomic DNA (both 5 µg/ml) and secretion of IFN1, IFN-α and TNF-α in the supernatants measured. (D) Dose-dependency of GAS DNA-mediated stimulation (12 hours) of IFN-α and TNF-α secretion. Data were pooled from 3 experiments done in triplicates and presented as mean ± SEM. * P<0.05 ** P<0.01.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375267&req=5

ppat-1002736-g001: Stimulation of murine macrophages by human and GAS DNA.(A–C) Bone marrow-derived mice macrophages (BMDMs) were stimulated with GAS and human genomic DNA (both 5 µg/ml) and secretion of IFN1, IFN-α and TNF-α in the supernatants measured. (D) Dose-dependency of GAS DNA-mediated stimulation (12 hours) of IFN-α and TNF-α secretion. Data were pooled from 3 experiments done in triplicates and presented as mean ± SEM. * P<0.05 ** P<0.01.
Mentions: DNA was purified from GAS strain 5448, representative of the globally disseminated hyperinvasive M1T1 clone, and incubated with murine bone marrow-derived macrophages (BMDMs). Cytokine release into the medium was used as readout for BMDM activation. GAS DNA induced time-dependent release of interferon type 1 (IFN-1), and specifically interferon-α (IFN-α), from the macrophages (Fig. 1A, B), peaking at 12 h of exposure. GAS DNA also induced BMDM TNF-α secretion, with maximal levels already detected after 6 h of incubation and remaining elevated for at least 24 h (Fig. 1C). Induction of IFN-α and TNF-α release by GAS DNA was also dose-dependent (Fig. 1D). In contrast, human DNA did not induce IFN-1 or TNF-α secretion from BMDMs (Fig. 1A–C). We did not detect specific induction of the cytokines IL-6, IL-1β, IL-10 or MIP-2 from BMDMs exposed to GAS DNA (data not shown).

Bottom Line: Similarly, in a murine necrotizing fasciitis model, IFN-α and TNF-α levels were significantly decreased in wild type mice infected with GAS expressing Sda1, whereas no such Sda1-dependent effect was seen in a TLR9-deficient background.Thus GAS Sda1 suppressed both the TLR9-mediated innate immune response and macrophage bactericidal activity.Our results demonstrate a novel mechanism of bacterial innate immune evasion based on autodegradation of CpG-rich DNA by a bacterial DNase.

View Article: PubMed Central - PubMed

Affiliation: Division of Infectious Diseases and Hospital Epidemiology, University Hospital Zurich, University of Zurich, Zurich, Switzerland.

ABSTRACT
Group A Streptococcus (GAS) has developed a broad arsenal of virulence factors that serve to circumvent host defense mechanisms. The virulence factor DNase Sda1 of the hyperinvasive M1T1 GAS clone degrades DNA-based neutrophil extracellular traps allowing GAS to escape extracellular killing. TLR9 is activated by unmethylated CpG-rich bacterial DNA and enhances innate immune resistance. We hypothesized that Sda1 degradation of bacterial DNA could alter TLR9-mediated recognition of GAS by host innate immune cells. We tested this hypothesis using a dual approach: loss and gain of function of DNase in isogenic GAS strains and presence and absence of TLR9 in the host. Either DNA degradation by Sda1 or host deficiency of TLR9 prevented GAS induced IFN-α and TNF-α secretion from murine macrophages and contributed to bacterial survival. Similarly, in a murine necrotizing fasciitis model, IFN-α and TNF-α levels were significantly decreased in wild type mice infected with GAS expressing Sda1, whereas no such Sda1-dependent effect was seen in a TLR9-deficient background. Thus GAS Sda1 suppressed both the TLR9-mediated innate immune response and macrophage bactericidal activity. Our results demonstrate a novel mechanism of bacterial innate immune evasion based on autodegradation of CpG-rich DNA by a bacterial DNase.

Show MeSH
Related in: MedlinePlus