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The AIM2 inflammasome is critical for innate immunity to Francisella tularensis.

Fernandes-Alnemri T, Yu JW, Juliana C, Solorzano L, Kang S, Wu J, Datta P, McCormick M, Huang L, McDermott E, Eisenlohr L, Landel CP, Alnemri ES - Nat. Immunol. (2010)

Bottom Line: Francisella tularensis, the causative agent of tularemia, infects host macrophages, which triggers production of the proinflammatory cytokines interleukin 1beta (IL-1beta) and IL-18.We elucidate here how host macrophages recognize F. tularensis and elicit this proinflammatory response.Caspase-1 activation, IL-1beta secretion and cell death were absent in Aim2(-/-) macrophages in response to F. tularensis infection or the presence of cytoplasmic DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania, USA.

ABSTRACT
Francisella tularensis, the causative agent of tularemia, infects host macrophages, which triggers production of the proinflammatory cytokines interleukin 1beta (IL-1beta) and IL-18. We elucidate here how host macrophages recognize F. tularensis and elicit this proinflammatory response. Using mice deficient in the DNA-sensing inflammasome component AIM2, we demonstrate here that AIM2 is required for sensing F. tularensis. AIM2-deficient mice were extremely susceptible to F. tularensis infection, with greater mortality and bacterial burden than that of wild-type mice. Caspase-1 activation, IL-1beta secretion and cell death were absent in Aim2(-/-) macrophages in response to F. tularensis infection or the presence of cytoplasmic DNA. Our study identifies AIM2 as a crucial sensor of F. tularensis infection and provides genetic proof of its critical role in host innate immunity to intracellular pathogens.

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Disruption of mouse Aim2 abolishes activation of the inflammasome by cytoplasmic DNA and vaccinia virus. (a) Immunoblot analysis of the expression of AIM2 in spleens and BMDMs from Aim2+/+ and Aim2−/− littermates (top) and in spleens from Aim2−/−, Aim2+/+ and Aim2+/− mice (bottom), assessed with an antibody specific for mouse AIM2. β-actin serves as a loading control. (b) Immunoblot (IB) analysis of mouse procaspase-1, caspase-1 (p20 subunit) and/or AIM2 in culture supernatants (Sup) and lysates (Lys) of Aim2+/+ and Aim2−/− BMDMs left untreated (None) or transfected with the synthetic DNA poly(dA:dT) or plasmid DNA (pcDNA), or treated for 5 h with LPS (500 ng/ml) followed by nigericin (2.5 µM) for 45 min (LPS + nig), assessed with monoclonal antibody to mouse caspase-1 p20 (anti-p20). (c) Release of LDH into culture supernatants of the BMDMs described in b, presented relative to the total cellular LDH content. *P < 0.05 and **P < 0.01, Aim2+/+ versus Aim2−/− (Student’s t-test). (d) Immunoblot analysis of mouse procaspase-1 and caspase-1 in culture supernatants and lysates of mouse Aim2−/− and Aim2+/+ BMDMs infected for 18 h with vaccinia virus (multiplicity of infection (MOI), above lanes). Data are representative of at least three experiments (mean and s.d. in c).
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Figure 1: Disruption of mouse Aim2 abolishes activation of the inflammasome by cytoplasmic DNA and vaccinia virus. (a) Immunoblot analysis of the expression of AIM2 in spleens and BMDMs from Aim2+/+ and Aim2−/− littermates (top) and in spleens from Aim2−/−, Aim2+/+ and Aim2+/− mice (bottom), assessed with an antibody specific for mouse AIM2. β-actin serves as a loading control. (b) Immunoblot (IB) analysis of mouse procaspase-1, caspase-1 (p20 subunit) and/or AIM2 in culture supernatants (Sup) and lysates (Lys) of Aim2+/+ and Aim2−/− BMDMs left untreated (None) or transfected with the synthetic DNA poly(dA:dT) or plasmid DNA (pcDNA), or treated for 5 h with LPS (500 ng/ml) followed by nigericin (2.5 µM) for 45 min (LPS + nig), assessed with monoclonal antibody to mouse caspase-1 p20 (anti-p20). (c) Release of LDH into culture supernatants of the BMDMs described in b, presented relative to the total cellular LDH content. *P < 0.05 and **P < 0.01, Aim2+/+ versus Aim2−/− (Student’s t-test). (d) Immunoblot analysis of mouse procaspase-1 and caspase-1 in culture supernatants and lysates of mouse Aim2−/− and Aim2+/+ BMDMs infected for 18 h with vaccinia virus (multiplicity of infection (MOI), above lanes). Data are representative of at least three experiments (mean and s.d. in c).

Mentions: To investigate the precise role of AIM2 in the host innate immune defense against dangerous cytosolic DNA produced by intracellular viral and microbial pathogens, we generated AIM2-deficient mice using the gene trap technology (Supplementary Fig. 1a,b) 12, 13. Immunoblot analysis of spleen and bone marrow samples from an Aim2−/− mouse and its Aim2+/+ littermate revealed the presence of full length AIM2 protein in the Aim2+/+ mouse but not in the Aim2−/− littermate (Fig. 1a, upper panels), indicating that the insertion of the gene trap vector in the Aim2 gene resulted in disruption of Aim2. Heterozygous Aim2+/− mice expressed reduced amount of AIM2 in the spleen compared to Aim2+/+ mice (Fig. 1a, lower panels), indicating that the expression of AIM2 is gene-dose dependent. All Aim2−/− mice had no obvious phenotypic abnormalities and were morphologically indistinguishable from their heterozygous or WT littermates (Supplementary Fig. 1c), indicating that AIM2 deficiency does not have any apparent adverse effects on mouse development.


The AIM2 inflammasome is critical for innate immunity to Francisella tularensis.

Fernandes-Alnemri T, Yu JW, Juliana C, Solorzano L, Kang S, Wu J, Datta P, McCormick M, Huang L, McDermott E, Eisenlohr L, Landel CP, Alnemri ES - Nat. Immunol. (2010)

Disruption of mouse Aim2 abolishes activation of the inflammasome by cytoplasmic DNA and vaccinia virus. (a) Immunoblot analysis of the expression of AIM2 in spleens and BMDMs from Aim2+/+ and Aim2−/− littermates (top) and in spleens from Aim2−/−, Aim2+/+ and Aim2+/− mice (bottom), assessed with an antibody specific for mouse AIM2. β-actin serves as a loading control. (b) Immunoblot (IB) analysis of mouse procaspase-1, caspase-1 (p20 subunit) and/or AIM2 in culture supernatants (Sup) and lysates (Lys) of Aim2+/+ and Aim2−/− BMDMs left untreated (None) or transfected with the synthetic DNA poly(dA:dT) or plasmid DNA (pcDNA), or treated for 5 h with LPS (500 ng/ml) followed by nigericin (2.5 µM) for 45 min (LPS + nig), assessed with monoclonal antibody to mouse caspase-1 p20 (anti-p20). (c) Release of LDH into culture supernatants of the BMDMs described in b, presented relative to the total cellular LDH content. *P < 0.05 and **P < 0.01, Aim2+/+ versus Aim2−/− (Student’s t-test). (d) Immunoblot analysis of mouse procaspase-1 and caspase-1 in culture supernatants and lysates of mouse Aim2−/− and Aim2+/+ BMDMs infected for 18 h with vaccinia virus (multiplicity of infection (MOI), above lanes). Data are representative of at least three experiments (mean and s.d. in c).
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Related In: Results  -  Collection

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Figure 1: Disruption of mouse Aim2 abolishes activation of the inflammasome by cytoplasmic DNA and vaccinia virus. (a) Immunoblot analysis of the expression of AIM2 in spleens and BMDMs from Aim2+/+ and Aim2−/− littermates (top) and in spleens from Aim2−/−, Aim2+/+ and Aim2+/− mice (bottom), assessed with an antibody specific for mouse AIM2. β-actin serves as a loading control. (b) Immunoblot (IB) analysis of mouse procaspase-1, caspase-1 (p20 subunit) and/or AIM2 in culture supernatants (Sup) and lysates (Lys) of Aim2+/+ and Aim2−/− BMDMs left untreated (None) or transfected with the synthetic DNA poly(dA:dT) or plasmid DNA (pcDNA), or treated for 5 h with LPS (500 ng/ml) followed by nigericin (2.5 µM) for 45 min (LPS + nig), assessed with monoclonal antibody to mouse caspase-1 p20 (anti-p20). (c) Release of LDH into culture supernatants of the BMDMs described in b, presented relative to the total cellular LDH content. *P < 0.05 and **P < 0.01, Aim2+/+ versus Aim2−/− (Student’s t-test). (d) Immunoblot analysis of mouse procaspase-1 and caspase-1 in culture supernatants and lysates of mouse Aim2−/− and Aim2+/+ BMDMs infected for 18 h with vaccinia virus (multiplicity of infection (MOI), above lanes). Data are representative of at least three experiments (mean and s.d. in c).
Mentions: To investigate the precise role of AIM2 in the host innate immune defense against dangerous cytosolic DNA produced by intracellular viral and microbial pathogens, we generated AIM2-deficient mice using the gene trap technology (Supplementary Fig. 1a,b) 12, 13. Immunoblot analysis of spleen and bone marrow samples from an Aim2−/− mouse and its Aim2+/+ littermate revealed the presence of full length AIM2 protein in the Aim2+/+ mouse but not in the Aim2−/− littermate (Fig. 1a, upper panels), indicating that the insertion of the gene trap vector in the Aim2 gene resulted in disruption of Aim2. Heterozygous Aim2+/− mice expressed reduced amount of AIM2 in the spleen compared to Aim2+/+ mice (Fig. 1a, lower panels), indicating that the expression of AIM2 is gene-dose dependent. All Aim2−/− mice had no obvious phenotypic abnormalities and were morphologically indistinguishable from their heterozygous or WT littermates (Supplementary Fig. 1c), indicating that AIM2 deficiency does not have any apparent adverse effects on mouse development.

Bottom Line: Francisella tularensis, the causative agent of tularemia, infects host macrophages, which triggers production of the proinflammatory cytokines interleukin 1beta (IL-1beta) and IL-18.We elucidate here how host macrophages recognize F. tularensis and elicit this proinflammatory response.Caspase-1 activation, IL-1beta secretion and cell death were absent in Aim2(-/-) macrophages in response to F. tularensis infection or the presence of cytoplasmic DNA.

View Article: PubMed Central - PubMed

Affiliation: Department of Biochemistry and Molecular Biology, Thomas Jefferson University, Philadelphia, Pennsylvania, USA.

ABSTRACT
Francisella tularensis, the causative agent of tularemia, infects host macrophages, which triggers production of the proinflammatory cytokines interleukin 1beta (IL-1beta) and IL-18. We elucidate here how host macrophages recognize F. tularensis and elicit this proinflammatory response. Using mice deficient in the DNA-sensing inflammasome component AIM2, we demonstrate here that AIM2 is required for sensing F. tularensis. AIM2-deficient mice were extremely susceptible to F. tularensis infection, with greater mortality and bacterial burden than that of wild-type mice. Caspase-1 activation, IL-1beta secretion and cell death were absent in Aim2(-/-) macrophages in response to F. tularensis infection or the presence of cytoplasmic DNA. Our study identifies AIM2 as a crucial sensor of F. tularensis infection and provides genetic proof of its critical role in host innate immunity to intracellular pathogens.

Show MeSH
Related in: MedlinePlus