Cyclic di-AMP in Mycobacterium tuberculosis

  • Yinlan Bai
  • Guangchun BaiEmail author


Mycobacterium tuberculosis (Mtb) is the etiologic agent of tuberculosis (TB), which is the leading cause of death worldwide by a single bacterial pathogen. This bacterium encodes a diadenylate cyclase, which is a homolog of Bacillus subtilis DNA integrity scanning protein A (DisA) and converts ATP into cyclic di-adenosine monophosphate (cyclic di-AMP). Mtb also possesses a DHH/DHHA1 family cyclic di-AMP phosphodiesterase, CnpB, which degrades cyclic di-AMP into AMP. Interestingly, elevating cyclic di-AMP levels by either overexpression of Mtb disA or deletion of cnpB in this pathogen results in significant virulence attenuation in a mouse pulmonary TB model. It has also been shown that cyclic di-AMP from Mtb activates autophagy and limits the growth of bacteria within infected cells. These findings indicate that cyclic di-AMP plays an important role in TB pathogenesis. Mtb exports cyclic di-AMP via an undefined mechanism, which induces a type I interferon response in a STING-dependent manner within the infected host. In contrast, the current live vaccine strain M. bovis BCG is unable to secrete cyclic di-AMP and is defective in inducing a type I interferon response. Thus, enabling the vaccine strain to induce type I interferon may provide better protection against infection of Mtb.


Mycobacterium tuberculosis Cyclic di-AMP DisA CnpB Pathogenesis Type I interferon Vaccine 


Conflict of Interest

The authors declare no conflict of interest.


  1. 1.
    Cernuschi T, Malvolti S, Nickels E, Friede M (2018) Bacillus Calmette-Guerin (BCG) vaccine: a global assessment of demand and supply balance. Vaccine 36:498–506PubMedPubMedCentralCrossRefGoogle Scholar
  2. 2.
    Delogu G, Sali M, Fadda G (2013) The biology of Mycobacterium tuberculosis infection. Mediterr J Hematol Infect Dis 5:e2013070PubMedPubMedCentralCrossRefGoogle Scholar
  3. 3.
    Forrellad MA, Klepp LI, Gioffre A, Sabio y Garcia J, Morbidoni HR, de la Paz Santangelo M, Cataldi AA, Bigi F (2013) Virulence factors of the Mycobacterium tuberculosis complex. Virulence 4:3–66PubMedPubMedCentralCrossRefGoogle Scholar
  4. 4.
    Weiss G, Schaible UE (2015) Macrophage defense mechanisms against intracellular bacteria. Immunol Rev 264:182–203PubMedPubMedCentralCrossRefGoogle Scholar
  5. 5.
    Etna MP, Giacomini E, Severa M, Coccia EM (2014) Pro- and anti-inflammatory cytokines in tuberculosis: a two-edged sword in TB pathogenesis. Semin Immunol 26:543–551PubMedCrossRefPubMedCentralGoogle Scholar
  6. 6.
    Takeuchi O, Akira S (2010) Pattern recognition receptors and inflammation. Cell 140:805–820PubMedCrossRefPubMedCentralGoogle Scholar
  7. 7.
    Hossain MM, Norazmi MN (2013) Pattern recognition receptors and cytokines in Mycobacterium tuberculosis infection—the double-edged sword? Biomed Res Int 2013:179174PubMedPubMedCentralCrossRefGoogle Scholar
  8. 8.
    Killick KE, Ni Cheallaigh C, O’Farrelly C, Hokamp K, MacHugh DE, Harris J (2013) Receptor-mediated recognition of mycobacterial pathogens. Cell Microbiol 15:1484–1495PubMedCrossRefPubMedCentralGoogle Scholar
  9. 9.
    Mortaz E, Adcock IM, Tabarsi P, Masjedi MR, Mansouri D, Velayati AA, Casanova JL, Barnes PJ (2015) Interaction of pattern recognition receptors with Mycobacterium tuberculosis. J Clin Immunol 35:1–10PubMedCrossRefPubMedCentralGoogle Scholar
  10. 10.
    McDonough KA, Rodriguez A (2011) The myriad roles of cyclic AMP in microbial pathogens: from signal to sword. Nat Rev Microbiol 10:27–38PubMedPubMedCentralCrossRefGoogle Scholar
  11. 11.
    Botsford JL, Harman JG (1992) Cyclic AMP in prokaryotes. Microbiol Rev 56:100–122PubMedPubMedCentralCrossRefGoogle Scholar
  12. 12.
    Agarwal N, Bishai WR (2009) cAMP signaling in Mycobacterium tuberculosis. Indian J Exp Biol 47:393–400PubMedPubMedCentralGoogle Scholar
  13. 13.
    Agarwal N, Lamichhane G, Gupta R, Nolan S, Bishai WR (2009) Cyclic AMP intoxication of macrophages by a Mycobacterium tuberculosis adenylate cyclase. Nature 460:98–102PubMedCrossRefPubMedCentralGoogle Scholar
  14. 14.
    Bai G, Schaak DD, McDonough KA (2009) cAMP levels within Mycobacterium tuberculosis and Mycobacterium bovis BCG increase upon infection of macrophages. FEMS Immunol Med Microbiol 55:68–73PubMedCrossRefPubMedCentralGoogle Scholar
  15. 15.
    Jenal U, Reinders A, Lori C (2017) Cyclic di-GMP: second messenger extraordinaire. Nat Rev Microbiol 15:271–284CrossRefGoogle Scholar
  16. 16.
    Romling U, Galperin MY, Gomelsky M (2013) Cyclic di-GMP: the first 25 years of a universal bacterial second messenger. Microbiol Mol Biol Rev 77:1–52PubMedPubMedCentralCrossRefGoogle Scholar
  17. 17.
    Kalia D, Merey G, Nakayama S, Zheng Y, Zhou J, Luo Y, Guo M, Roembke BT, Sintim HO (2013) Nucleotide, c-di-GMP, c-di-AMP, cGMP, cAMP, (p)ppGpp signaling in bacteria and implications in pathogenesis. Chem Soc Rev 42:305–341CrossRefGoogle Scholar
  18. 18.
    Gupta K, Kumar P, Chatterji D (2010) Identification, activity and disulfide connectivity of C-di-GMP regulating proteins in Mycobacterium tuberculosis. PLoS One 5:e15072PubMedPubMedCentralCrossRefGoogle Scholar
  19. 19.
    Kumar M, Chatterji D (2008) Cyclic di-GMP: a second messenger required for long-term survival, but not for biofilm formation, in Mycobacterium smegmatis. Microbiology 154:2942–2955CrossRefGoogle Scholar
  20. 20.
    Bharati BK, Sharma IM, Kasetty S, Kumar M, Mukherjee R, Chatterji D (2012) A full-length bifunctional protein involved in c-di-GMP turnover is required for long-term survival under nutrient starvation in Mycobacterium smegmatis. Microbiology 158:1415–1427CrossRefGoogle Scholar
  21. 21.
    Witte G, Hartung S, Buttner K, Hopfner KP (2008) Structural biochemistry of a bacterial checkpoint protein reveals diadenylate cyclase activity regulated by DNA recombination intermediates. Mol Cell 30:167–178PubMedPubMedCentralCrossRefGoogle Scholar
  22. 22.
    Fahmi T, Port GC, Cho KH (2017) c-di-AMP: an essential molecule in the signaling pathways that regulate the viability and virulence of gram-positive bacteria. Genes (Basel) 8. PubMedCentralCrossRefGoogle Scholar
  23. 23.
    Devaux L, Kaminski PA, Trieu-Cuot P, Firon A (2018) Cyclic di-AMP in host-pathogen interactions. Curr Opin Microbiol 41:21–28CrossRefGoogle Scholar
  24. 24.
    Corrigan RM, Grundling A (2013) Cyclic di-AMP: another second messenger enters the fray. Nat Rev Microbiol 11:513–524PubMedPubMedCentralCrossRefGoogle Scholar
  25. 25.
    Bai Y, Yang J, Eisele LE, Underwood AJ, Koestler BJ, Waters CM, Metzger DW, Bai G (2013) Two DHH subfamily 1 proteins in Streptococcus pneumoniae possess cyclic di-AMP phosphodiesterase activity and affect bacterial growth and virulence. J Bacteriol 195:5123–5132PubMedPubMedCentralCrossRefGoogle Scholar
  26. 26.
    Corrigan RM, Abbott JC, Burhenne H, Kaever V, Grundling A (2011) c-di-AMP is a new second messenger in Staphylococcus aureus with a role in controlling cell size and envelope stress. PLoS Pathog 7:e1002217PubMedPubMedCentralCrossRefGoogle Scholar
  27. 27.
    Witte CE, Whiteley AT, Burke TP, Sauer JD, Portnoy DA, Woodward JJ (2013) Cyclic di-AMP is critical for Listeria monocytogenes growth, cell wall homeostasis, and establishment of infection. MBio 4:e00282–e00213PubMedPubMedCentralCrossRefGoogle Scholar
  28. 28.
    Luo Y, Helmann JD (2012) Analysis of the role of Bacillus subtilis sigma(M) in β-lactam resistance reveals an essential role for c-di-AMP in peptidoglycan homeostasis. Mol Microbiol 83:623–639PubMedPubMedCentralCrossRefGoogle Scholar
  29. 29.
    Mehne FM, Gunka K, Eilers H, Herzberg C, Kaever V, Stulke J (2013) Cyclic di-AMP homeostasis in Bacillus subtilis: both lack and high level accumulation of the nucleotide are detrimental for cell growth. J Biol Chem 288:2004–2017PubMedPubMedCentralCrossRefGoogle Scholar
  30. 30.
    Zhang Y, Yang J, Bai G (2018) Cyclic di-AMP-mediated interaction between Mycobacterium tuberculosis ΔcnpB and macrophages implicates a novel strategy for improving BCG vaccination. Pathog Dis 76.
  31. 31.
    Bai Y, Yang J, Zhou X, Ding X, Eisele LE, Bai G (2012) Mycobacterium tuberculosis Rv3586 (DacA) is a diadenylate cyclase that converts ATP or ADP into c-di-AMP. PLoS One 7:e35206PubMedPubMedCentralCrossRefGoogle Scholar
  32. 32.
    Dey B, Dey RJ, Cheung LS, Pokkali S, Guo H, Lee JH, Bishai WR (2015) A bacterial cyclic dinucleotide activates the cytosolic surveillance pathway and mediates innate resistance to tuberculosis. Nat Med 21:401–406PubMedPubMedCentralCrossRefGoogle Scholar
  33. 33.
    Yang J, Bai Y, Zhang Y, Gabrielle VD, Jin L, Bai G (2014) Deletion of the cyclic di-AMP phosphodiesterase gene (cnpB) in Mycobacterium tuberculosis leads to reduced virulence in a mouse model of infection. Mol Microbiol 93:65–79PubMedPubMedCentralCrossRefGoogle Scholar
  34. 34.
    Tang Q, Luo Y, Zheng C, Yin K, Ali MK, Li X, He J (2015) Functional Analysis of a c-di-AMP-specific phosphodiesterase MsPDE from Mycobacterium smegmatis. Int J Biol Sci 11:813–824PubMedPubMedCentralCrossRefGoogle Scholar
  35. 35.
    Rao F, See RY, Zhang D, Toh DC, Ji Q, Liang ZX (2010) YybT is a signaling protein that contains a cyclic dinucleotide phosphodiesterase domain and a GGDEF domain with ATPase activity. J Biol Chem 285:473–482PubMedPubMedCentralCrossRefGoogle Scholar
  36. 36.
    Dey RJ, Dey B, Zheng Y, Cheung LS, Zhou J, Sayre D, Kumar P, Guo H, Lamichhane G, Sintim HO, Bishai WR (2017) Inhibition of innate immune cytosolic surveillance by an M. tuberculosis phosphodiesterase. Nat Chem Biol 13:210–217CrossRefGoogle Scholar
  37. 37.
    He Q, Wang F, Liu S, Zhu D, Cong H, Gao F, Li B, Wang H, Lin Z, Liao J, Gu L (2016) Structural and biochemical insight into the mechanism of Rv2837c from Mycobacterium tuberculosis as a c-di-NMP phosphodiesterase. J Biol Chem 291:3668–3681CrossRefGoogle Scholar
  38. 38.
    Manikandan K, Sabareesh V, Singh N, Saigal K, Mechold U, Sinha KM (2014) Two-step synthesis and hydrolysis of cyclic di-AMP in Mycobacterium tuberculosis. PLoS One 9:e86096PubMedPubMedCentralCrossRefGoogle Scholar
  39. 39.
    Postic G, Danchin A, Mechold U (2012) Characterization of NrnA homologs from Mycobacterium tuberculosis and Mycoplasma pneumoniae. RNA 18:155–165PubMedPubMedCentralCrossRefGoogle Scholar
  40. 40.
    Zhang Y, Yang J, Bai G (2018) Regulation of the CRISPR-associated genes by Rv2837c (CnpB) via an Orn-like activity in TB complex mycobacteria. J Bacteriol.
  41. 41.
    Gundlach J, Herzberg C, Kaever V, Gunka K, Hoffmann T, Weiss M, Gibhardt J, Thurmer A, Hertel D, Daniel R, Bremer E, Commichau FM, Stulke J (2017) Control of potassium homeostasis is an essential function of the second messenger cyclic di-AMP in Bacillus subtilis. Sci Signal 10. PubMedPubMedCentralCrossRefGoogle Scholar
  42. 42.
    Whiteley AT, Pollock AJ, Portnoy DA (2015) The PAMP c-di-AMP is essential for Listeria monocytogenes growth in rich but not minimal media due to a toxic increase in (p)ppGpp. [corrected]. Cell Host Microbe 17:788–798PubMedPubMedCentralCrossRefGoogle Scholar
  43. 43.
    Zeden MS, Schuster CF, Bowman L, Zhong Q, Williams HD, Grundling A (2018) Cyclic di-adenosine monophosphate (c-di-AMP) is required for osmotic regulation in Staphylococcus aureus but dispensable for viability in anaerobic conditions. J Biol Chem 293:3180–3200PubMedPubMedCentralCrossRefGoogle Scholar
  44. 44.
    Zhang L, He ZG (2013) Radiation-sensitive gene A (RadA) targets DisA, DNA integrity scanning protein A, to negatively affect cyclic Di-AMP synthesis activity in Mycobacterium smegmatis. J Biol Chem 288:22426–22436PubMedPubMedCentralCrossRefGoogle Scholar
  45. 45.
    Bejerano-Sagie M, Oppenheimer-Shaanan Y, Berlatzky I, Rouvinski A, Meyerovich M, Ben-Yehuda S (2006) A checkpoint protein that scans the chromosome for damage at the start of sporulation in Bacillus subtilis. Cell 125:679–690PubMedPubMedCentralCrossRefGoogle Scholar
  46. 46.
    Oppenheimer-Shaanan Y, Wexselblatt E, Katzhendler J, Yavin E, Ben-Yehuda S (2011) c-di-AMP reports DNA integrity during sporulation in Bacillus subtilis. EMBO Rep 12:594–601PubMedPubMedCentralCrossRefGoogle Scholar
  47. 47.
    Zhang L, Li W, He ZG (2013) DarR, a TetR-like transcriptional factor, is a cyclic di-AMP-responsive repressor in Mycobacterium smegmatis. J Biol Chem 288:3085–3096CrossRefGoogle Scholar
  48. 48.
    Cho KH, Kang SO (2013) Streptococcus pyogenes c-di-AMP phosphodiesterase, GdpP, influences SpeB processing and virulence. PLoS One 8:e69425PubMedPubMedCentralCrossRefGoogle Scholar
  49. 49.
    Du B, Ji W, An H, Shi Y, Huang Q, Cheng Y, Fu Q, Wang H, Yan Y, Sun J (2014) Functional analysis of c-di-AMP phosphodiesterase, GdpP, in Streptococcus suis serotype 2. Microbiol Res 169:749–758PubMedPubMedCentralCrossRefGoogle Scholar
  50. 50.
    Huynh TN, Luo S, Pensinger D, Sauer JD, Tong L, Woodward JJ (2015) An HD-domain phosphodiesterase mediates cooperative hydrolysis of c-di-AMP to affect bacterial growth and virulence. Proc Natl Acad Sci USA 112:E747–E756PubMedPubMedCentralCrossRefGoogle Scholar
  51. 51.
    Peng X, Zhang Y, Bai G, Zhou X, Wu H (2016) Cyclic di-AMP mediates biofilm formation. Mol Microbiol 99:945–959PubMedPubMedCentralCrossRefGoogle Scholar
  52. 52.
    Ye M, Zhang JJ, Fang X, Lawlis GB, Troxell B, Zhou Y, Gomelsky M, Lou Y, Yang XF (2014) DhhP, a cyclic di-AMP phosphodiesterase of Borrelia burgdorferi, is essential for cell growth and virulence. Infect Immun 82:1840–1849PubMedPubMedCentralCrossRefGoogle Scholar
  53. 53.
    Sauer JD, Sotelo-Troha K, von Moltke J, Monroe KM, Rae CS, Brubaker SW, Hyodo M, Hayakawa Y, Woodward JJ, Portnoy DA, Vance RE (2011) The N-ethyl-N-nitrosourea-induced Goldenticket mouse mutant reveals an essential function of STING in the in vivo interferon response to Listeria monocytogenes and cyclic dinucleotides. Infect Immun 79:688–694PubMedCrossRefPubMedCentralGoogle Scholar
  54. 54.
    Schwartz KT, Carleton JD, Quillin SJ, Rollins SD, Portnoy DA, Leber JH (2012) Hyperinduction of host beta interferon by a Listeria monocytogenes strain naturally overexpressing the multidrug efflux pump MdrT. Infect Immun 80:1537–1545PubMedPubMedCentralCrossRefGoogle Scholar
  55. 55.
    Woodward JJ, Iavarone AT, Portnoy DA (2010) c-di-AMP secreted by intracellular Listeria monocytogenes activates a host type I interferon response. Science 328:1703–1705PubMedPubMedCentralCrossRefGoogle Scholar
  56. 56.
    Yamamoto T, Hara H, Tsuchiya K, Sakai S, Fang R, Matsuura M, Nomura T, Sato F, Mitsuyama M, Kawamura I (2012) Listeria monocytogenes strain-specific impairment of the TetR regulator underlies the drastic increase in cyclic di-AMP secretion and beta interferon-inducing ability. Infect Immun 80:2323–2332PubMedPubMedCentralCrossRefGoogle Scholar
  57. 57.
    Archer KA, Durack J, Portnoy DA (2014) STING-dependent type I IFN production inhibits cell-mediated immunity to Listeria monocytogenes. PLoS Pathog 10:e1003861PubMedPubMedCentralCrossRefGoogle Scholar
  58. 58.
    Collins AC, Cai H, Li T, Franco LH, Li XD, Nair VR, Scharn CR, Stamm CE, Levine B, Chen ZJ, Shiloh MU (2015) Cyclic GMP-AMP synthase is an innate immune DNA sensor for Mycobacterium tuberculosis. Cell Host Microbe 17:820–828PubMedPubMedCentralCrossRefGoogle Scholar
  59. 59.
    Wassermann R, Gulen MF, Sala C, Perin SG, Lou Y, Rybniker J, Schmid-Burgk JL, Schmidt T, Hornung V, Cole ST, Ablasser A (2015) Mycobacterium tuberculosis differentially activates cGAS- and inflammasome-dependent intracellular immune responses through ESX-1. Cell Host Microbe 17:799–810PubMedCrossRefPubMedCentralGoogle Scholar
  60. 60.
    Watson RO, Bell SL, MacDuff DA, Kimmey JM, Diner EJ, Olivas J, Vance RE, Stallings CL, Virgin HW, Cox JS (2015) The cytosolic sensor cGAS detects Mycobacterium tuberculosis DNA to induce type I interferons and activate autophagy. Cell Host Microbe 17:811–819PubMedPubMedCentralCrossRefGoogle Scholar
  61. 61.
    Kaplan Zeevi M, Shafir NS, Shaham S, Friedman S, Sigal N, Nir Paz R, Boneca IG, Herskovits AA (2013) Listeria monocytogenes multidrug resistance transporters and cyclic di-AMP, which contribute to type I interferon induction, play a role in cell wall stress. J Bacteriol 195:5250–5261PubMedPubMedCentralCrossRefGoogle Scholar
  62. 62.
    Tadmor K, Pozniak Y, Burg Golani T, Lobel L, Brenner M, Sigal N, Herskovits AA (2014) Listeria monocytogenes MDR transporters are involved in LTA synthesis and triggering of innate immunity during infection. Front Cell Infect Microbiol 4:16PubMedPubMedCentralCrossRefGoogle Scholar
  63. 63.
    Giacomini E, Remoli ME, Gafa V, Pardini M, Fattorini L, Coccia EM (2009) IFN-β improves BCG immunogenicity by acting on DC maturation. J Leukoc Biol 85:462–468PubMedCrossRefPubMedCentralGoogle Scholar
  64. 64.
    Guerrero GG, Rangel-Moreno J, Islas-Trujillo S, Rojas-Espinosa O (2015) Successive intramuscular boosting with IFN-α protects Mycobacterium bovis BCG-vaccinated mice against M. lepraemurium infection. Biomed Res Int 2015:414027PubMedPubMedCentralCrossRefGoogle Scholar
  65. 65.
    Groschel MI, Sayes F, Shin SJ, Frigui W, Pawlik A, Orgeur M, Canetti R, Honore N, Simeone R, van der Werf TS, Bitter W, Cho SN, Majlessi L, Brosch R (2017) Recombinant BCG expressing ESX-1 of Mycobacterium marinum combines low virulence with cytosolic immune signaling and improved TB protection. Cell Rep 18:2752–2765PubMedCrossRefPubMedCentralGoogle Scholar
  66. 66.
    Manzanillo PS, Shiloh MU, Portnoy DA, Cox JS (2012) Mycobacterium tuberculosis activates the DNA-dependent cytosolic surveillance pathway within macrophages. Cell Host Microbe 11:469–480PubMedPubMedCentralCrossRefGoogle Scholar
  67. 67.
    Behr MA, Wilson MA, Gill WP, Salamon H, Schoolnik GK, Rane S, Small PM (1999) Comparative genomics of BCG vaccines by whole-genome DNA microarray. Science 284:1520–1523PubMedCrossRefPubMedCentralGoogle Scholar
  68. 68.
    Conrad WH, Osman MM, Shanahan JK, Chu F, Takaki KK, Cameron J, Hopkinson-Woolley D, Brosch R, Ramakrishnan L (2017) Mycobacterial ESX-1 secretion system mediates host cell lysis through bacterium contact-dependent gross membrane disruptions. Proc Natl Acad Sci USA 114:1371–1376PubMedCrossRefPubMedCentralGoogle Scholar
  69. 69.
    Stanley SA, Johndrow JE, Manzanillo P, Cox JS (2007) The Type I IFN response to infection with Mycobacterium tuberculosis requires ESX-1-mediated secretion and contributes to pathogenesis. J Immunol 178:3143–3152PubMedCrossRefPubMedCentralGoogle Scholar

Copyright information

© Springer Nature Switzerland AG 2020

Authors and Affiliations

  1. 1.Department of Microbiology, College of Basic Medical SciencesAir Force Military Medical UniversityXi’anChina
  2. 2.Department of Immunology and Microbial DiseaseAlbany Medical CollegeAlbanyUSA

Personalised recommendations