Skip to main content
NCBI home page
Protein & Cell logoLink to Protein & Cell
. 2011 Aug 6;2(7):564–572. doi: 10.1007/s13238-011-1078-2

AMPD3 is involved in anthrax LeTx-induced macrophage cell death

Sangun Lee 1, Yanhai Wang 1,2,✉, Sung Ouk Kim 1, Jiahuai Han 2,✉
PMCID: PMC4875241  PMID: 21822801

Abstract

The responses of macrophages to Bacillus anthracis infection are important for the survival of the host, since macrophages are required for the germination of B. anthracis spores in lymph nodes, and macrophage death exacerbates anthrax lethal toxin (LeTx)-induced organ collapse. To elucidate the mechanism of macrophage cell death induced by LeTx, we performed a genetic screen to search for genes associated with LeTx-induced macrophage cell death. RAW264.7 cells, a macrophage-like cell line sensitive to LeTx-induced death, were randomly mutated and LeTx-resistant mutant clones were selected. AMP deaminase 3 (AMPD3), an enzyme that converts AMP to IMP, was identified to be mutated in one of the resistant clones. The requirement of AMPD3 in LeTxinduced cell death of RAW 264.7 cells was confirmed by the restoration of LeTx sensitivity with ectopic reconstitution of AMPD3 expression. AMPD3 deficiency does not affect LeTx entering cells and the cleavage of mitogen-activated protein kinase kinase (MKK) by lethal factor inside cells, but does impair an unknown downstream event that is linked to cell death. Our data provides new information regarding LeTx-induced macrophage death and suggests that there is a key regulatory site downstream of or parallel to MKK cleavage that controls the cell death in LeTx-treated macrophages.

Keywords: AMP deaminase 3, anthrax lethal toxin, macrophage, cell death

Contributor Information

Yanhai Wang, Email: wangyh@xmu.edu.cn.

Jiahuai Han, Email: jhan@scripps.edu.

References

  1. Banks D.J., Barnajian M., Maldonado-Arocho F.J., Sanchez A.M., Bradley K.A. Anthrax toxin receptor 2 mediates Bacillus anthracis killing of macrophages following spore challenge. Cell Microbiol. 2005;7:1173–1185. doi: 10.1111/j.1462-5822.2005.00545.x. [DOI] [PubMed] [Google Scholar]
  2. Bazan J.F., Koch-Nolte F. Sequence and structural links between distant ADP-ribosyltransferase families. Adv Exp Med Biol. 1997;419:99–107. doi: 10.1007/978-1-4419-8632-0_12. [DOI] [PubMed] [Google Scholar]
  3. Duesbery N.S., Webb C.P., Leppla S.H., Gordon V.M., Klimpel K. R., Copeland T.D., et al. Proteolytic inactivation of MAPkinase-kinase by anthrax lethal factor. Science. 1998;280:734–737. doi: 10.1126/science.280.5364.734. [DOI] [PubMed] [Google Scholar]
  4. Friedlander A.M. Macrophages are sensitive to anthrax lethal toxin through an acid-dependent process. J Biol Chem. 1986;261:7123–7126. [PubMed] [Google Scholar]
  5. Friedlander A.M., Bhatnagar R., Leppla S.H., Johnson L., Singh Y. Characterization of macrophage sensitivity and resistance to anthrax lethal toxin. Infect Immun. 1993;61:245–252. doi: 10.1128/iai.61.1.245-252.1993. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Guidi-Rontani C., Levy M., Ohayon H., Mock M. Fate of germinated Bacillus anthracis spores in primary murine macrophages. Mol Microbiol. 2001;42:931–938. doi: 10.1046/j.1365-2958.2001.02695.x. [DOI] [PubMed] [Google Scholar]
  7. Kim S.O., Ha S.D., Lee S., Stanton S., Beutler B., Han J. Mutagenesis by retroviral insertion in chemical mutagengenerated quasi-haploid mammalian cells. Biotechniques. 2007;42:493–501. doi: 10.2144/000112390. [DOI] [PubMed] [Google Scholar]
  8. Kim S.O., Jing Q., Hoebe K., Beutler B., Duesbery N.S., Han J. Sensitizing anthrax lethal toxin-resistant macrophages to lethal toxin-induced killing by tumor necrosis factor-alpha. J Biol Chem. 2003;278:7413–7421. doi: 10.1074/jbc.M209279200. [DOI] [PubMed] [Google Scholar]
  9. McAllister R.D., Singh Y., Du Bois W.D., Potter M., Boehm T., Meeker N.D., Fillmore P.D., Anderson L.M., Poynter M.E., Teuscher C. Susceptibility to anthrax lethal toxin is controlled by three linked quantitative trait loci. Am J Pathol. 2003;163:1735–1741. doi: 10.1016/S0002-9440(10)63532-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Moayeri M., Haines D., Young H.A., Leppla S.H. Bacillus anthracis lethal toxin induces TNF-independent hypoxiamediated toxicity in mice. J Clin Invest. 2003;112:670–682. doi: 10.1172/JCI17991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Moayeri M., Leppla S.H. Cellular and systemic effects of anthrax lethal toxin and edema toxin. Mol Aspects Med. 2009;30:439–455. doi: 10.1016/j.mam.2009.07.003. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Moayeri M., Martinez N.W., Wiggins J., Young H.A., Leppla S. H. Mouse susceptibility to anthrax lethal toxin is influenced by genetic factors in addition to those controlling macrophage sensitivity. Infect Immun. 2004;72:4439–4447. doi: 10.1128/IAI.72.8.4439-4447.2004. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Moayeri M., Webster J.I., Wiggins J.F., Leppla S.H., Sternberg E.M. Endocrine perturbation increases susceptibility of mice to anthrax lethal toxin. Infect Immun. 2005;73:4238–4244. doi: 10.1128/IAI.73.7.4238-4244.2005. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Morisaki H., Morisaki T. AMPD genes and urate metabolism. Nippon Rinsho. 2008;66:771–777. [PubMed] [Google Scholar]
  15. Muehlbauer S.M., Evering T.H., Bonuccelli G., Squires R.C., Ashton A.W., Porcelli S.A., Lisanti M.P., Brojatsch J. Anthrax lethal toxin kills macrophages in a strain-specific manner by apoptosis or caspase-1-mediated necrosis. Cell Cycle. 2007;6:758–766. doi: 10.4161/cc.6.6.3991. [DOI] [PubMed] [Google Scholar]
  16. Park J.M., Greten F.R., Li Z.W., Karin M. Macrophage apoptosis by anthrax lethal factor through p38 MAP kinase inhibition. Science. 2002;297:2048–2051. doi: 10.1126/science.1073163. [DOI] [PubMed] [Google Scholar]
  17. Pellizzari R., Guidi-Rontani C., Vitale G., Mock M., Montecucco C. Anthrax lethal factor cleaves MKK3 in macrophages and inhibits the LPS/IFNgamma-induced release of NO and TNFalpha. FEBS Lett. 1999;462:199–204. doi: 10.1016/S0014-5793(99)01502-1. [DOI] [PubMed] [Google Scholar]
  18. Scobie H.M., Young J.A. Interactions between anthrax toxin receptors and protective antigen. Curr Opin Microbiol. 2005;8:106–112. doi: 10.1016/j.mib.2004.12.005. [DOI] [PubMed] [Google Scholar]
  19. Smith H. Discovery of the anthrax toxin: the beginning of in vivo studies on pathogenic bacteria. Trends Microbiol. 2000;8:199–200. doi: 10.1016/S0966-842X(00)01755-8. [DOI] [PubMed] [Google Scholar]
  20. Smith H. Discovery of the anthrax toxin: the beginning of studies of virulence determinants regulated in vivo. Int J Med Microbiol. 2002;291:411–417. doi: 10.1078/1438-4221-00147. [DOI] [PubMed] [Google Scholar]
  21. Smith H., Keppie J. Observations on experimental anthrax: demonstration of a specific lethal factor produced in vivo by Bacillus anthracis. Nature. 1954;173:869–870. doi: 10.1038/173869a0. [DOI] [PubMed] [Google Scholar]
  22. Vitale G., Bernardi L., Napolitani G., Mock M., Montecucco C. Susceptibility of mitogen-activated protein kinase kinase family members to proteolysis by anthrax lethal factor. Biochem J. 2000;352:739–745. doi: 10.1042/bj3520739. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Vitale G., Pellizzari R., Recchi C., Napolitani G., Mock M., Montecucco C. Anthrax lethal factor cleaves the Nterminus of MAPKKs and induces tyrosine/threonine phosphorylation of MAPKs in cultured macrophages. Biochem Biophys Res Commun. 1998;248:706–711. doi: 10.1006/bbrc.1998.9040. [DOI] [PubMed] [Google Scholar]
  24. Wang X., Ono K., Kim S.O., Kravchenko V., Lin S.C., Han J. Metaxin is required for tumor necrosis factorinduced cell death. EMBO Rep. 2001;2:628–633. doi: 10.1093/embo-reports/kve135. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Protein & Cell are provided here courtesy of Oxford University Press

RESOURCES