1887

Abstract

The ability of to persist in its human host despite extensive chemotherapy is thought to be based on subpopulations of non-replicating phenotypically drug-resistant bacilli. To study the non-growing pathogen, culture models that generate quiescent organisms by either oxygen depletion in nutrient-rich medium (Wayne model) or nutrient deprivation in oxygen-rich medium (Loebel model) have been developed. In contrast to the energy metabolism of Wayne bacilli, little is known about Loebel bacilli. Here we analysed under nutrient-starvation conditions. Upon shifting to the non-replicating state the pathogen maintained a fivefold reduced but constant intracellular ATP level. Chemical probing of the FF ATP synthase demonstrated the importance of this enzyme for ATP homeostasis and viability of the nutrient-starved organism. Surprisingly, the specific ATP synthase inhibitor TMC207 did not affect viability and only moderately reduced the intracellular ATP level of nutrient-starved organisms. Depletion of oxygen killed Loebel bacilli, whereas death was prevented by nitrate, suggesting that respiration and an exogenous electron acceptor are required for maintaining viability. Nutrient-starved bacilli lacking the glyoxylate shunt enzyme isocitrate lyase failed to reduce their intracellular ATP level and died, thus establishing a link between ATP control and intermediary metabolism. We conclude that reduction of the ATP level might be an important step in the adaptation of to non-growing survival.

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2010-01-01
2019-11-17
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References

  1. Amaral, L., Kristiansen, J. E., Abebe, L. S. & Millett, W. ( 1996; ). Inhibition of the respiration of multi-drug resistant clinical isolates of Mycobacterium tuberculosis by thioridazine: potential use for initial therapy of freshly diagnosed tuberculosis. J Antimicrob Chemother 38, 1049–1053.[CrossRef]
    [Google Scholar]
  2. Andries, K., Verhasselt, P., Guillemont, J., Göhlmann, H. W., Neefs, J. M., Winkler, H., Van Gestel, J., Timmerman, P., Zhu, M. & other authors ( 2005; ). A diarylquinoline drug active on the ATP synthase of Mycobacterium tuberculosis. Science 307, 223–227.[CrossRef]
    [Google Scholar]
  3. Bardarov, S., Bardarov, S., Jr, Pavelka, M. S., Jr, Sambandamurthy, V., Larsen, M., Tufariello, J., Chan, J., Hatfull, G. & Jacobs, W. R., Jr ( 2002; ). Specialized transduction: an efficient method for generating marked and unmarked targeted gene disruptions in Mycobacterium tuberculosis, M. bovis BCG and M. smegmatis. Microbiology 148, 3007–3017.
    [Google Scholar]
  4. Betts, J. C., Lukey, P. T., Robb, L. C., McAdam, R. A. & Duncan, K. ( 2002; ). Evaluation of a nutrient starvation model of Mycobacterium tuberculosis persistence by gene and protein expression profiling. Mol Microbiol 43, 717–731.[CrossRef]
    [Google Scholar]
  5. Boshoff, H. I. & Barry, C. E., III ( 2005; ). Tuberculosis – metabolism and respiration in the absence of growth. Nat Rev Microbiol 3, 70–80.[CrossRef]
    [Google Scholar]
  6. Cattell, K. J., Lindop, C. R., Knight, I. G. & Beechey, R. B. ( 1971; ). The identification of the site of action of N,N′-dicyclohexylcarbodi-imide as a proteolipid in mitochondrial membranes. Biochem J 125, 169–177.
    [Google Scholar]
  7. Dahl, J. L., Kraus, C. N., Boshoff, H. I., Doan, B., Foley, K., Avarbock, D., Kaplan, G., Mizrahi, V., Rubin, H. & Barry, C. E., III ( 2003; ). The role of RelMtb-mediated adaptation to stationary phase in long-term persistence of Mycobacterium tuberculosis in mice. Proc Natl Acad Sci U S A 100, 10026–10031.[CrossRef]
    [Google Scholar]
  8. Dick, T. ( 2001; ). Dormant tubercle bacilli: the key to more effective TB chemotherapy? J Antimicrob Chemother 47, 117–118.[CrossRef]
    [Google Scholar]
  9. Fenhalls, G., Stevens, L., Moses, L., Bezuidenhout, J., Betts, J. C., Helden, P. van, Lukey, P. T. & Duncan, K. ( 2002; ). In situ detection of Mycobacterium tuberculosis transcripts in human lung granulomas reveals differential gene expression in necrotic lesions. Infect Immun 70, 6330–6338.[CrossRef]
    [Google Scholar]
  10. Gordhan, B. G., Smith, D. A., Kana, B. D., Bancroft, G. & Mizrahi, V. ( 2006; ). The carbon starvation-inducible genes Rv2557 and Rv2558 of Mycobacterium tuberculosis are not required for long-term survival under carbon starvation and for virulence in SCID mice. Tuberculosis (Edinb) 86, 430–437.[CrossRef]
    [Google Scholar]
  11. Harries, A. D. & Dye, C. ( 2006; ). Tuberculosis. Ann Trop Med Parasitol 100, 415–431.
    [Google Scholar]
  12. Hinds, J., Mahenthiralingam, E., Kempsell, K. E., Duncan, K., Stokes, R. W., Parish, T. & Stoker, N. G. ( 1999; ). Enhanced gene replacement in mycobacteria. Microbiology 145, 519–527.[CrossRef]
    [Google Scholar]
  13. Honer Zu Bentrup, K., Miczak, A., Swenson, D. L. & Russell, D. G. ( 1999; ). Characterization of activity and expression of isocitrate lyase in Mycobacterium avium and Mycobacterium tuberculosis. J Bacteriol 181, 7161–7167.
    [Google Scholar]
  14. Koul, A., Dendouga, N., Vergauwen, K., Molenberghs, B., Vranckx, L., Willebrords, R., Ristic, Z., Lill, H., Dorange, I. & other authors ( 2007; ). Diarylquinolines target subunit c of mycobacterial ATP synthase. Nat Chem Biol 3, 323–324.[CrossRef]
    [Google Scholar]
  15. Koul, A., Vranckx, L., Dendouga, N., Balemans, W., Van den Wyngaert, I., Vergauwen, K., Göhlmann, H. W., Willebrords, R., Poncelet, A. & other authors ( 2008; ). Diarylquinolines are bactericidal for dormant mycobacteria as a result of disturbed ATP homeostasis. J Biol Chem 283, 25273–25280.[CrossRef]
    [Google Scholar]
  16. Lee, M. H., Pascopella, L., Jacobs, W. R., Jr & Hatfull, G. F. ( 1991; ). Site-specific integration of mycobacteriophage L5: integration-proficient vectors for Mycobacterium smegmatis, Mycobacterium tuberculosis, and bacille Calmette-Guerin. Proc Natl Acad Sci U S A 88, 3111–3115.[CrossRef]
    [Google Scholar]
  17. Lim, A., Eleuterio, M., Hutter, B., Murugasu-Oei, B. & Dick, T. ( 1999; ). Oxygen depletion-induced dormancy in Mycobacterium bovis BCG. J Bacteriol 181, 2252–2256.
    [Google Scholar]
  18. Loebel, R. O., Shorr, E. & Richardson, H. B. ( 1933a; ). The influence of adverse conditions upon the respiratory metabolism and growth of human tubercle bacilli. J Bacteriol 26, 167–200.
    [Google Scholar]
  19. Loebel, R. O., Shorr, E. & Richardson, H. B. ( 1933b; ). The influence of foodstuffs upon the respiratory metabolism and growth of human tubercle bacilli. J Bacteriol 26, 139–166.
    [Google Scholar]
  20. McKinney, J. D., Honer zu Bentrup, K., Munoz-Elias, E. J., Miczak, A., Chen, B., Chan, W. T., Swenson, D., Sacchettini, J. C., Jacobs, W. R., Jr & Russell, D. G. ( 2000; ). Persistence of Mycobacterium tuberculosis in macrophages and mice requires the glyoxylate shunt enzyme isocitrate lyase. Nature 406, 735–738.[CrossRef]
    [Google Scholar]
  21. Nyka, W. ( 1974; ). Studies on the effect of starvation on mycobacteria. Infect Immun 9, 843–850.
    [Google Scholar]
  22. Ovchinnikov, Y. A.., Abdulaev, N. G. & Modyanov, N. N. ( 1982; ). Structural basis of proton-translocating protein function. Annu Rev Biophys Bioeng 11, 445–463.[CrossRef]
    [Google Scholar]
  23. Parish, T. & Stoker, N. G. ( 2000; ). Use of a flexible cassette method to generate a double unmarked Mycobacterium tuberculosis tlyA plcABC mutant by gene replacement. Microbiology 146, 1969–1975.
    [Google Scholar]
  24. Primm, T. P., Andersen, S. J., Mizrahi, V., Avarbock, D., Rubin, H. & Barry, C. E., III ( 2000; ). The stringent response of Mycobacterium tuberculosis is required for long-term survival. J Bacteriol 182, 4889–4898.[CrossRef]
    [Google Scholar]
  25. Rao, S. P., Alonso, S., Rand, L., Dick, T. & Pethe, K. ( 2008; ). The protonmotive force is required for maintaining ATP homeostasis and viability of hypoxic, nonreplicating Mycobacterium tuberculosis. Proc Natl Acad Sci U S A 105, 11945–11950.[CrossRef]
    [Google Scholar]
  26. Stover, C. K., de la Cruz, V. F., Fuerst, T. R., Burlein, J. E., Benson, L. A., Bennett, L. T., Bansal, G. P., Young, J. F. & Lee, M. H. ( 1991; ). New use of BCG for recombinant vaccines. Nature 351, 456–460.[CrossRef]
    [Google Scholar]
  27. Teh, J. S., Yano, T. & Rubin, H. ( 2007; ). Type II NADH : menaquinone oxidoreductase of Mycobacterium tuberculosis. Infect Disord Drug Targets 7, 169–181.[CrossRef]
    [Google Scholar]
  28. Via, L. E., Lin, P. L., Ray, S. M., Carrillo, J., Allen, S. S., Eum, S. Y., Taylor, K., Klein, E., Manjunatha, U. & other authors ( 2008; ). Tuberculous granulomas are hypoxic in guinea pigs, rabbits, and nonhuman primates. Infect Immun 76, 2333–2340.[CrossRef]
    [Google Scholar]
  29. Wayne, L. G. & Hayes, L. G. ( 1996; ). An in vitro model for sequential study of shiftdown of Mycobacterium tuberculosis through two stages of nonreplicating persistence. Infect Immun 64, 2062–2069.
    [Google Scholar]
  30. Wayne, L. G. & Sohaskey, C. D. ( 2001; ). Nonreplicating persistence of Mycobacterium tuberculosis. Annu Rev Microbiol 55, 139–163.[CrossRef]
    [Google Scholar]
  31. Weinstein, E. A., Yano, T., Li, L. S. & other authors ( 2005; ). Inhibitors of type II NADH : menaquinone oxidoreductase represent a class of antitubercular drugs. Proc Natl Acad Sci U S A 102, 4548–4553.[CrossRef]
    [Google Scholar]
  32. Williams, K. J. & Duncan, K. ( 2007; ). Current strategies for identifying and validating targets for new treatment-shortening drugs for TB. Curr Mol Med 7, 297–307.[CrossRef]
    [Google Scholar]
  33. Xie, Z., Siddiqi, N. & Rubin, E. J. ( 2005; ). Differential antibiotic susceptibilities of starved Mycobacterium tuberculosis isolates. Antimicrob Agents Chemother 49, 4778–4780.[CrossRef]
    [Google Scholar]
  34. Yano, T., Li, L. S., Weinstein, E., Teh, J. S. & Rubin, H. ( 2006; ). Steady-state kinetics and inhibitory action of antitubercular phenothiazines on Mycobacterium tuberculosis type-II NADH-menaquinone oxidoreductase (NDH-2). J Biol Chem 281, 11456–11463.[CrossRef]
    [Google Scholar]
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NADH dehydrogenase-2 but not 1 is required for survival of nutrient-starved non-growing tubercle bacilli [ PDF] (21 kb)



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