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. 1992 Nov;3(11):1269–1277. doi: 10.1091/mbc.3.11.1269

Biochemical and physiological changes induced by anthrax lethal toxin in J774 macrophage-like cells.

P C Hanna 1, S Kochi 1, R J Collier 1
PMCID: PMC275693  PMID: 1457831

Abstract

Experiments were performed to probe the mechanism by which Bacillus anthracis Lethal Toxin (LeTx) causes lysis of J774 macrophage-like cells. After incubation of cells with saturating concentrations of the toxin, two categories of effects were found, which were distinguishable on the basis of chronology, Ca(2+)-dependence, and sensitivity to osmolarity. The earliest events (category I), beginning 45 min postchallenge, were an increase in permeability to 22Na and 86Rb and a rapid conversion of ATP to ADP and AMP. Later events (category II) included alterations in membrane permeability to 45Ca, 51Cr, 36Cl, 35SO4, 3H-amino acids, and 3H-uridine, beginning at 60 min; inhibition of macromolecular synthesis, leakage of cellular lactate dehydrogenase and onset of gross morphological changes, at approximately 75 min; and cell lysis, beginning at 90 min. Category II events exhibited an absolute requirement for extracellular Ca2+ and were blocked by addition of 0.3 M sucrose to the medium, whereas category I events were attenuated, but not blocked, by either of these conditions. On the other hand, both ATP depletion and the category II events were blocked in osmotically stabilized medium that was also isoionic for Na+ and K+. This suggests that permeabilization of the plasma membrane to monovalent cations and water may be the earliest of the physiological changes described here. The resulting influx of Na+ and efflux of K+ would be expected to cause depletion of ATP, via increased activity of the Na+/K+ pump. Subsequently the influx of Ca2+, induced by depletion of ATP, imbalances in monovalent cautions, and/or more dramatic changes in permeability due to influx of water, would be expected to trigger widespread changes leading ultimately to cytolysis.

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Selected References

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  1. Bhakdi S., Tranum-Jensen J. Damage to cell membranes by pore-forming bacterial cytolysins. Prog Allergy. 1988;40:1–43. [PubMed] [Google Scholar]
  2. Bhatnagar R., Singh Y., Leppla S. H., Friedlander A. M. Calcium is required for the expression of anthrax lethal toxin activity in the macrophagelike cell line J774A.1. Infect Immun. 1989 Jul;57(7):2107–2114. doi: 10.1128/iai.57.7.2107-2114.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Blaustein R. O., Koehler T. M., Collier R. J., Finkelstein A. Anthrax toxin: channel-forming activity of protective antigen in planar phospholipid bilayers. Proc Natl Acad Sci U S A. 1989 Apr;86(7):2209–2213. doi: 10.1073/pnas.86.7.2209. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Boobis A. R., Fawthrop D. J., Davies D. S. Mechanisms of cell death. Trends Pharmacol Sci. 1989 Jul;10(7):275–280. doi: 10.1016/0165-6147(89)90027-8. [DOI] [PubMed] [Google Scholar]
  5. Carafoli E. Intracellular calcium homeostasis. Annu Rev Biochem. 1987;56:395–433. doi: 10.1146/annurev.bi.56.070187.002143. [DOI] [PubMed] [Google Scholar]
  6. Cataldi A., Labruyère E., Mock M. Construction and characterization of a protective antigen-deficient Bacillus anthracis strain. Mol Microbiol. 1990 Jul;4(7):1111–1117. doi: 10.1111/j.1365-2958.1990.tb00685.x. [DOI] [PubMed] [Google Scholar]
  7. Chang M. P., Bramhall J., Graves S., Bonavida B., Wisnieski B. J. Internucleosomal DNA cleavage precedes diphtheria toxin-induced cytolysis. Evidence that cell lysis is not a simple consequence of translation inhibition. J Biol Chem. 1989 Sep 15;264(26):15261–15267. [PubMed] [Google Scholar]
  8. DePetrillo P. B., Swift R. M., Ambroise C., Abernethy D. R. High-performance liquid chromatographic determination of 3',5'-cyclic adenosine monophosphate in human platelets. J Chromatogr. 1990 May 18;527(2):421–427. doi: 10.1016/s0378-4347(00)82126-x. [DOI] [PubMed] [Google Scholar]
  9. Escuyer V., Collier R. J. Anthrax protective antigen interacts with a specific receptor on the surface of CHO-K1 cells. Infect Immun. 1991 Oct;59(10):3381–3386. doi: 10.1128/iai.59.10.3381-3386.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Friedlander A. M. Macrophages are sensitive to anthrax lethal toxin through an acid-dependent process. J Biol Chem. 1986 Jun 5;261(16):7123–7126. [PubMed] [Google Scholar]
  11. Harris R. W., Sims P. J., Tweten R. K. Evidence that Clostridium perfringens theta-toxin induces colloid-osmotic lysis of erythrocytes. Infect Immun. 1991 Jul;59(7):2499–2501. doi: 10.1128/iai.59.7.2499-2501.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Iacono-Connors L. C., Welkos S. L., Ivins B. E., Dalrymple J. M. Protection against anthrax with recombinant virus-expressed protective antigen in experimental animals. Infect Immun. 1991 Jun;59(6):1961–1965. doi: 10.1128/iai.59.6.1961-1965.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Koehler T. M., Collier R. J. Anthrax toxin protective antigen: low-pH-induced hydrophobicity and channel formation in liposomes. Mol Microbiol. 1991 Jun;5(6):1501–1506. doi: 10.1111/j.1365-2958.1991.tb00796.x. [DOI] [PubMed] [Google Scholar]
  14. Leppla S. H. Anthrax toxin edema factor: a bacterial adenylate cyclase that increases cyclic AMP concentrations of eukaryotic cells. Proc Natl Acad Sci U S A. 1982 May;79(10):3162–3166. doi: 10.1073/pnas.79.10.3162. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Leppla S. H. Production and purification of anthrax toxin. Methods Enzymol. 1988;165:103–116. doi: 10.1016/s0076-6879(88)65019-1. [DOI] [PubMed] [Google Scholar]
  16. McClane B. A., Hanna P. C., Wnek A. P. Clostridium perfringens enterotoxin. Microb Pathog. 1988 May;4(5):317–323. doi: 10.1016/0882-4010(88)90059-9. [DOI] [PubMed] [Google Scholar]
  17. McClane B. A., McDonel J. L. Protective effects of osmotic stabilizers on morphological and permeability alterations induced in Vero cells by Clostridium perfringens enterotoxin. Biochim Biophys Acta. 1981 Mar 6;641(2):401–409. doi: 10.1016/0005-2736(81)90496-x. [DOI] [PubMed] [Google Scholar]
  18. McClane B. A., McDonel J. L. The effects of Clostridium perfringens enterotoxin on morphology, viability, and macromolecular synthesis in Vero cells. J Cell Physiol. 1979 May;99(2):191–200. doi: 10.1002/jcp.1040990205. [DOI] [PubMed] [Google Scholar]
  19. McClane B. A., Wnek A. P., Hulkower K. I., Hanna P. C. Divalent cation involvement in the action of Clostridium perfringens type A enterotoxin. Early events in enterotoxin action are divalent cation-independent. J Biol Chem. 1988 Feb 15;263(5):2423–2435. [PubMed] [Google Scholar]
  20. Orrenius S., McConkey D. J., Bellomo G., Nicotera P. Role of Ca2+ in toxic cell killing. Trends Pharmacol Sci. 1989 Jul;10(7):281–285. doi: 10.1016/0165-6147(89)90029-1. [DOI] [PubMed] [Google Scholar]
  21. Pezard C., Berche P., Mock M. Contribution of individual toxin components to virulence of Bacillus anthracis. Infect Immun. 1991 Oct;59(10):3472–3477. doi: 10.1128/iai.59.10.3472-3477.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Thelestam M., Möllby R. Sensitive assay for detection of toxin-induced damage to the cytoplasmic membrane of human diploid fibroblasts. Infect Immun. 1975 Aug;12(2):225–232. doi: 10.1128/iai.12.2.225-232.1975. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Welch R. A. Pore-forming cytolysins of gram-negative bacteria. Mol Microbiol. 1991 Mar;5(3):521–528. doi: 10.1111/j.1365-2958.1991.tb00723.x. [DOI] [PubMed] [Google Scholar]
  24. Welkos S. L., Friedlander A. M. Comparative safety and efficacy against Bacillus anthracis of protective antigen and live vaccines in mice. Microb Pathog. 1988 Aug;5(2):127–139. doi: 10.1016/0882-4010(88)90015-0. [DOI] [PubMed] [Google Scholar]
  25. Wyllie A. H., Kerr J. F., Currie A. R. Cell death: the significance of apoptosis. Int Rev Cytol. 1980;68:251–306. doi: 10.1016/s0074-7696(08)62312-8. [DOI] [PubMed] [Google Scholar]

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