Anthrax lethal factor cleaves MKK3 in macrophages and inhibits the LPS/IFNγ‐induced release of NO and TNFα

The lethal toxin of Bacillus anthracis consists of two proteins, PA and LF, which together induce lethal effects in animals and cause macrophage lysis. LF is a zinc‐endopeptidase which cleaves two mitogen‐activated proten kinase kinases (MAPKKs), Mek1 and Mek2, within the cytosol. Here, we show that also MKK3, another dual‐specificity kinase that phosphorylates and activates p38 MAP kinase, is cleaved by LF in macrophages. No direct correlation between LF‐induced cell death and cleavage of these MAPKKs was found in macrophage cell lines and primary peritoneal cells exhibiting different sensitivity to LF. However, we present the first evidence that sublytic doses of LF cleave Meks and cause a substantial reduction in the production of NO and tumour necrosis factor‐α induced by lipopolysaccharide/interferonγ. We suggest that this effect of LF is relevant during the first stages of B. anthracis infection, when a reduction of the inflammatory response would permit growth and diffusion of the bacterium.

[1]  J. Heesemann,et al.  Yersinia enterocolitica Promotes Deactivation of Macrophage Mitogen-activated Protein Kinases Extracellular Signal-regulated Kinase-1/2, p38, and c-Jun NH2-terminal Kinase , 1997, The Journal of Biological Chemistry.

[2]  W. Fiers,et al.  The p38/RK mitogen‐activated protein kinase pathway regulates interleukin‐6 synthesis response to tumor necrosis factor. , 1996, The EMBO journal.

[3]  C. Widmann,et al.  Mitogen-activated protein kinase: conservation of a three-kinase module from yeast to human. , 1999, Physiological reviews.

[4]  R. Collier,et al.  Protective antigen‐binding domain of anthrax lethal factor mediates translocation of a heterologous protein fused to its amino‐ or carboxy‐terminus , 1995, Molecular microbiology.

[5]  J. Watters,et al.  Ltx1, a mouse locus that influences the susceptibility of macrophages to cytolysis caused by intoxication with Bacillus anthracis lethal factor, maps to chromosome 11 , 1998, Molecular microbiology.

[6]  P. Ricciardi-Castagnoli,et al.  Mouse macrophage clones immortalized by retroviruses are functionally heterogeneous. , 1991, Proceedings of the National Academy of Sciences of the United States of America.

[7]  M. Mcdaniel,et al.  Nitric Oxide Regulates Interleukin 1 Bioactivity Released from Murine Macrophages* , 1996, The Journal of Biological Chemistry.

[8]  D. Acosta,et al.  On the role of macrophages in anthrax. , 1993, Proceedings of the National Academy of Sciences of the United States of America.

[9]  P. Hanna,et al.  Biochemical and physiological changes induced by anthrax lethal toxin in J774 macrophage-like cells. , 1992, Molecular biology of the cell.

[10]  S. Welkos,et al.  Pathogenesis and genetic control of resistance to the Sterne strain of Bacillus anthracis. , 1988, Microbial pathogenesis.

[11]  R. Hill,et al.  The Biogenetic Anatomy of Vitamin B6 , 1996, The Journal of Biological Chemistry.

[12]  R. Rappuoli,et al.  Guidebook to protein toxins and their use in cell biology , 1997 .

[13]  M. Mock,et al.  Construction of Bacillus anthracis mutant strains producing a single toxin component. , 1993, Journal of general microbiology.

[14]  R. Bhatnagar,et al.  Protein synthesis is required for expression of anthrax lethal toxin cytotoxicity , 1994, Infection and immunity.

[15]  D. Pappin,et al.  Cdc2 Kinase Directly Phosphorylates the cis-Golgi Matrix Protein GM130 and Is Required for Golgi Fragmentation in Mitosis , 1998, Cell.

[16]  G. Wright,et al.  Anthrax toxin blocks priming of neutrophils by lipopolysaccharide and by muramyl dipeptide , 1986, The Journal of experimental medicine.

[17]  Jerry L. Adams,et al.  A protein kinase involved in the regulation of inflammatory cytokine biosynthesis , 1994, Nature.

[18]  M. Mock,et al.  Germination of Bacillus anthracis spores within alveolar macrophages , 1999, Molecular microbiology.

[19]  A. Friedlander,et al.  Macrophages are sensitive to anthrax lethal toxin through an acid-dependent process. , 1986, The Journal of biological chemistry.

[20]  U. Rapp,et al.  Different Mitogen-activated Protein Kinase Signaling Pathways Cooperate to Regulate Tumor Necrosis Factor α Gene Expression in T Lymphocytes* , 1999, The Journal of Biological Chemistry.

[21]  S. Leppla,et al.  Characterization of macrophage sensitivity and resistance to anthrax lethal toxin , 1993, Infection and immunity.

[22]  C. Nathan,et al.  Release of reactive nitrogen intermediates and reactive oxygen intermediates from mouse peritoneal macrophages. Comparison of activating cytokines and evidence for independent production. , 1988, Journal of immunology.

[23]  M. Mock,et al.  The effects of pH on the interaction of anthrax toxin lethal and edema factors with phospholipid vesicles. , 1994, Biochemistry.

[24]  C. Nathan,et al.  Nitric oxide and macrophage function. , 1997, Annual review of immunology.

[25]  M. Mock,et al.  Anthrax lethal factor cleaves the N-terminus of MAPKKs and induces tyrosine/threonine phosphorylation of MAPKs in cultured macrophages. , 1998, Biochemical and biophysical research communications.

[26]  M. Mock,et al.  The cytotoxic activity of Bacillus anthracis lethal factor is inhibited by leukotriene A4 hydrolase and metallopeptidase inhibitors. , 1996, The Biochemical journal.

[27]  M. Mock,et al.  The vacuolar ATPase proton pump is required for the cytotoxicity of Bacillus anthracis lethal toxin , 1996, FEBS letters.

[28]  S. Welkos,et al.  Differences in susceptibility of inbred mice to Bacillus anthracis , 1986, Infection and immunity.

[29]  S. Leppla,et al.  Anthrax toxin edema factor: a bacterial adenylate cyclase that increases cyclic AMP concentrations of eukaryotic cells. , 1982, Proceedings of the National Academy of Sciences of the United States of America.

[30]  H. Smith,et al.  The three factors of anthrax toxin: their immunogenicity and lack of demonstrable enzymic activity. , 1963, Journal of general microbiology.

[31]  M. Hansen,et al.  Re-examination and further development of a precise and rapid dye method for measuring cell growth/cell kill. , 1989, Journal of immunological methods.

[32]  S. Leppla,et al.  Internalization and processing of Bacillus anthracis lethal toxin by toxin-sensitive and -resistant cells. , 1989, The Journal of biological chemistry.

[33]  B. Bloom,et al.  Role of Macrophage Oxidative Burst in the Action of Anthrax Lethal Toxin , 1994, Molecular medicine.

[34]  K D Paull,et al.  Proteolytic inactivation of MAP-kinase-kinase by anthrax lethal factor. , 1998, Science.

[35]  M. Mock,et al.  Contribution of individual toxin components to virulence of Bacillus anthracis , 1991, Infection and immunity.