Mode of Action of Cry Toxins from Bacillus thuringiensis and Resistance Mechanisms

Living reference work entry
Part of the Toxinology book series (TOXI)

Abstract

Bacillus thuringiensis (Bt) insecticidal Cry toxins have been shown to be effective in controlling insect pests either in spray products or expressed in transgenic crops. All Cry toxins are expressed as protoxins that undergo proteolytic processing in the insect gut releasing the activated toxin. It has been shown that activated toxin binds to different insect protein molecules in gut cells leading to oligomerization, membrane insertion, and pore formation. However, it was recently shown that not only the activated toxin is able to specifically interact with receptors, since Cry1A protoxins bind gut receptor molecules leading also to oligomerization, membrane insertion, and pore formation. The final pores induced by protoxin or by activated toxin have different characteristics, suggesting dual mode of action of Cry proteins. In addition it was shown that different Cry1A resistant populations from different insect species are significantly more susceptible to Cry1A protoxins than to Cry1A activated toxins, supporting that Cry1A proteins may undergo two toxic pathways one involving protoxin binding to receptors and another involving the binding of activated Cry toxins to gut receptor molecules. Here the authors will revise this dual mode of action of Cry proteins and discuss implications of the dual mode of action of Cry proteins for insect pest management in transgenic plants.

Keywords

Bacillus thuringiensis Cry toxins Insect resistance Pore formation Mode of action 

References

  1. Anilkumar KJ, Rodrigo-Simón A, Ferré J, Pusztai-Carey M, Sivasupramaniam S, Moar WJ. Production and characterization of Bacillus thuringiensis Cry1Ac-resistant cotton bollworm Helicoverpa zea (Boddie). Appl Environ Microbiol. 2008;74:462–9.CrossRefPubMedGoogle Scholar
  2. Boonserm P, Davis P, Ellar DJ, Li J. Crystal structure of the mosquito-larvicidal toxin Cry4Ba and its biological implications. J Mol Biol. 2005;348:363–82.CrossRefPubMedGoogle Scholar
  3. Bravo A, Gómez I, Conde J, Muñoz-Garay C, Sánchez J, Zhuang M, Gill SS, Soberón M. Oligomerization triggers binding of a Bacillus thuringiensis Cry1Ab pore-forming toxin to aminopeptidase N receptor leading to insertion into membrane microdomains. Biochim Biophys Acta. 2004;1667:38–46.CrossRefPubMedGoogle Scholar
  4. Bravo A, Martinez-de-Castro DL, Sánchez-Quintana J, Cantón PE, Mendoza G, Gómez I, Pacheco S, García-Gómez BI, Onofre J, Soberón M. Mechanism of action of Bacillus thuringiensis insecticidal toxins and their use in the control of insect pests. In: Alouf JE, Ladant D, Popoff MR, editors. Comprehensive sourcebook of bacterial protein toxins. 4ath ed. Boston: Acad Press; 2015.Google Scholar
  5. Chakroun M, Banylus N, Bel Y, Escriche B, Ferre J. Bacterial vegetative insecticidal proteins (Vip) from entomopathogenic bacteria. Microbiol Mol Biol Rev. 2016;80(2):329–50.CrossRefPubMedGoogle Scholar
  6. Crickmore N, Baum J, Bravo A, Lereclus D, Narva K, Sampson K, Schnepf E, Sun M, Zeigler DR. Bacillus thuringiensis toxin nomenclature [Internet]. 2016. Available from http://www.btnomenclature.info/
  7. de Maagd R, Bravo A, Crickmore N. How Bacillus thuringiensis has evolved specific toxins to colonize the insect world. TIG. 2001;17:193–9.CrossRefPubMedGoogle Scholar
  8. Evdokimov A, Moshiri F, Sturman EJ, Rydel TJ, Zheng M, Seale JW, Franklin S. Structure of the full-length insecticidal protein Cry1Ac reveals intriguing details of toxin packaging into in vivo formed crystals. Protein Sci. 2014;23:1491–7.CrossRefPubMedPubMedCentralGoogle Scholar
  9. Fabrick JA, Tabashnik BE. Binding of Bacillus thuringiensis toxin Cry1Ac to multiple sites of cadherin in pink bollworm. Insect Biochem Mol Biol. 2007;37:97–106.CrossRefPubMedGoogle Scholar
  10. Fabrick JA, Mathew LG, Tabashnik BE, Li X. Insertion of an intact CR1 retrotransposon in a cadherin gene linked with Bt resistance in the pink bollworm, Pectinophora gossypiella. Insect Mol Biol. 2011;20:651–65.CrossRefPubMedGoogle Scholar
  11. Flores-Escobar B, Rodríguez-Magadan H, Bravo A, Soberón M, Gómez I. Manduca sexta aminopeptidase-n and alkaline phosphatase have a differential role in the mode of action of Cry1Aa, Cry1Ab and Cry1Ac toxins from Bacillus thuringiensis. Appl Environ Microbiol. 2013;79:4543–50.CrossRefPubMedPubMedCentralGoogle Scholar
  12. Gahan LJ, Gould F, Heckel DG. Identification of a gene associated with Bt resistance in Heliothis virescens. Science. 2001;293:857–60.CrossRefPubMedGoogle Scholar
  13. Gassman AJ, Petzold-Maxwell JL, Keweshan RS, Dunbar MW. Field-evolved resistance to Bt maize by western corn rootworm. PLoS One. 2013;6(7):e22629.CrossRefGoogle Scholar
  14. Girard F, Vachon V, Prefontaine G, Marceau L, Larouche G, Vincent C, Schwartz J-L, Masson L, Laprade R. Cysteine scanning mutagenesis of alpha 4 a putative pore forming helix of the Bacillus thuringiensis insecticidal toxin Cry1Aa. Appl Environ Microbiol. 2008;74:2565–72.CrossRefPubMedPubMedCentralGoogle Scholar
  15. Gómez I, Sánchez J, Miranda R, Bravo A, Soberón M. Cadherin-like receptor binding facilitates proteolytic cleavage of helix α-1 in domain I and oligomer pre-pore formation of Bacillus thuringiensis Cry1Ab toxin. FEBS Lett. 2002;513:242–6.CrossRefPubMedGoogle Scholar
  16. Gómez I, Arenas I, Benitez I, Miranda-Ríos J, Becerril B, Grande AR, Almagro JC, Bravo A, Soberón M. Specific epitopes of Domains II and III of Bacillus thuringiensis Cry1Ab toxin involved in the sequential interaction with cadherin and aminopeptidase-N receptors in Manduca sexta. J Biol Chem. 2006;281:34032–9.CrossRefPubMedGoogle Scholar
  17. Gómez I, Sanchez J, Muñoz-Garay C, Matus V, Gill SS, Soberón M, Bravo A. Bacillus thuringiensis Cry1A toxins are versatile-proteins with multiple modes of action: two distinct pre-pores are involved in toxicity. Biochem J. 2014;459:383–96.CrossRefPubMedPubMedCentralGoogle Scholar
  18. Heckel D. Roles of ABC proteins in the mechanism and management of Bt resistance. In: Soberón M, Gao Y, Bravo A, editors. Bt resistance – characterization and strategies for GM crops expressing Bacillus thuringiensis toxins. CABI; Wallingford, Oxfordshire, 2015.Google Scholar
  19. Herrero S, Gechev T, Bakker PL, Moar WJ, de Maagd RA. Bacillus thuringiensis Cry1Ca-resistant Spodoptera exigua lacks expression of one of four aminopeptidase N genes. BMC Genomics. 2005;24:6–96.Google Scholar
  20. Hua G, Jurat-Fuentes JL, Adang MJ. Fluorescent based assay establish Manduca sexta Bt-R1 cadherin as receptor for multiple Bacillus thuringiensis Cry1A toxins in Drosophila S2 cells. Insect Biochem Mol Biol. 2004;34:193–202.CrossRefPubMedGoogle Scholar
  21. Hui F, Scheib U, Hu Y, Sommer RJ, Aroian RV, Ghosh P. Structure and glycolipid binding properties of the nematicidal protein Cry5B. Biochemistry. 2012;51:9911–21.CrossRefPubMedPubMedCentralGoogle Scholar
  22. James C. Global status of commercialized biotech/GM Crops: 2015, ISAAA briefs, vol. 51. Ithaca: ISAAA; 2015.Google Scholar
  23. Jiménez-Juárez A, Muñoz-Garay C, Gómez I, Saab-Rincon G, Damian-Almazo JY, Gill SS, Soberón M, Bravo A. Bacillus thuringiensis Cry1Ab mutants affecting oligomer formation are non-toxic to Manduca sexta larvae. J Biol Chem. 2007;282:21222–9.CrossRefPubMedGoogle Scholar
  24. Jurat Fuentes JL, Adang MJ. The Heliothis virescens cadherin protein expressed in Drosophila S2 cells functions as a receptor for Bacillus thuringiensis Cry1A but not Cry1Fa toxins. Biochemistry. 2006;45:9688–95.CrossRefPubMedGoogle Scholar
  25. Jurat-Fuentes JL, Karumbaiah L, Jakka SRK, Ning C, Liu C, Wu K, Jackson J, Gould F, Blanco C, Portilla M, Perera O, Adang M. Reduced levels of membrane-bound alkaline phosphatase are common to lepidopteran strains resistant to Cry toxins from Bacillus thuringiensis. PLoS One. 2011;6:e17606.CrossRefPubMedPubMedCentralGoogle Scholar
  26. Koul B, Yadav R, Sanyal I, Amla DV. Comparative performance of modified full-length and truncated Bacillus thuringiensis-cry1Ac genes in transgenic tomato. Springerplus. 2015;4:203.CrossRefPubMedPubMedCentralGoogle Scholar
  27. Kwa MSG, de Maagd RA, Stiekema WJ, Vlak JM, Bosch D. Toxicity and binding properties of the Bacillus thuringiensis delta-endotoxin Cry1C to cultured insect cells. J Invertebr Pathol. 1998;71:121–7.CrossRefPubMedGoogle Scholar
  28. Liu K, Zheng B, Hong H, Jiang C, Peng R, Peng J, Yu Z, Zheng J, Yang H. Characterization of cultured insect cells selected by Bacillus thuringiensis crystal toxins. In Vitro Cell Dev Biol Anim. 2004;40:312–7.CrossRefPubMedGoogle Scholar
  29. Lorence A, Darszon A, Díaz C, Liévano A, Quintero R, Bravo A. Delta-endotoxins induce cation channels in Spodoptera frugiperda brush border membrane in suspension and in planar lipid bilayers. FEBS Lett. 1995;360:353–6.Google Scholar
  30. Martin FG, Wolfersberger MG. Bacillus thuringiensis delta-endotoxin and larval Manduca sexta midgut brush border membrane vesicles act synergistically to cause very large increases in the conductance of planar lipid bilayers. J Exp Biol. 1995;198:91–6.PubMedGoogle Scholar
  31. Monnerat R, Martins E, Macedo C, Queiroz P, Praça L, Soares CM, Moreira H, Grisi I, Silva J, Soberón M, Bravo A. Evidence of field-evolved resistance of Spodoptera frugiperda to Bt corn expressing Cry1F in Brazil that is still sensitive to modified Bt toxins. PLoS One. 2015;10:e0119544.CrossRefPubMedPubMedCentralGoogle Scholar
  32. Morin S, Biggs RW, Shriver L, Ellers-Kirk C, Higginson D, Holley D, Gahan LJ, Heckel DG, Carriere Y, Dennehy TJ, Brown JK, Tabashnik BE. Three cadherin alleles associated with resistance to Bacillus thuringiensis in pink bollworm. Proc Natl Acad Sci U S A. 2003;100:5004–9.CrossRefPubMedPubMedCentralGoogle Scholar
  33. Morse RJ, Yamamoto T, Stroud RM. Structure of Cry2Aa suggests an unexpected receptor binding epitope. Structure. 2001;9:409–17.CrossRefPubMedGoogle Scholar
  34. Muñoz-Garay C, Rodríguez-Almazán C, Aguilar JN, Portugal L, Gómez I, Saab-Rincon G, Soberón M, Bravo A. Oligomerization of Cry11Aa from Bacillus thuringiensis has an important role in toxicity against Aedes aegypti. Appl Environ Microbiol. 2009;75:7548–50.CrossRefPubMedPubMedCentralGoogle Scholar
  35. Pacheco S, Gomez I, Arenas I, Saab-Rincon G, Rodriguez-Almazan C, Gill SS, Bravo A, Soberón M. Domain II loop 3 of Bacillus thuringiensis Cry1Ab toxin is involved in a “ping-pong” binding mechanism with Manduca sexta aminopeptidase-N and cadherin receptors. J Biol Chem. 2009;284:32750–7.CrossRefPubMedPubMedCentralGoogle Scholar
  36. Pardo-López L, Soberón M, Bravo A. Bacillus thuringiensis insecticidal 3-domain Cry toxins: mode of action, insect resistance and consequences for crop protection. FEMS Microbiol Rev. 2013;37:3–22.CrossRefPubMedGoogle Scholar
  37. Peyronnet O, Vachon V, Schwartz JL, Laprade R. Ion Channels in planar lipid bilayers by the Bacillus thuringiensis toxin Cry1Aa in the presence of gypsy moth (Lymantria dispar) brush border membrane. J Membr Biol. 2001;184:45–54.CrossRefPubMedGoogle Scholar
  38. Pigott CR, Ellar DJ. Role of receptors in Bacillus thuringiensis crystal toxin activity. Microbiol Mol Biol Rev. 2007;71:255–81.CrossRefPubMedPubMedCentralGoogle Scholar
  39. Qaim M, Zilberman D. Yield effects of genetically modified crops in developing countries. Science. 2003;299:900–2.CrossRefPubMedGoogle Scholar
  40. Rajagopal R, Sivakumar S, Agrawai N, Malhotra P, Bhatnagar RK. Silencing of midgut aminopeptidase N of Spodoptera litura by double-stranded RNA establishes its role as Bacillus thuringiensis toxin receptor. J Biol Chem. 2002;277:46849–51.CrossRefPubMedGoogle Scholar
  41. Rodríguez-Almazán C, Zavala LE, Muñoz-Garay C, Jiménez-Juárez N, Pacheco S, Masson L, Soberón M, Bravo A. Dominant negative mutants of Bacillus thuringiensis Cry1Ab toxin function as anti-toxins: demonstration of the role of oligomerization in toxicity. PLoS One. 2009;4:e5545.CrossRefPubMedPubMedCentralGoogle Scholar
  42. Sanahuja G, Banakar R, Twyman RM, Capell T, Christou P. Bacillus thuringiensis: a century of research development and commercial applications. Plant Biotechnol J. 2011;9:283–300.CrossRefPubMedGoogle Scholar
  43. Sangadala S, Walters FS, English LH, Adang MJ. A mixture of Manduca sexta aminopeptidase and phosphatase enhances Bacillus thuringiensis insecticidal Cry1Ac toxin binding and86Rb+-K+ efflux in vitro. J Biol Chem. 1994;269:10088–92.PubMedGoogle Scholar
  44. Schwartz JL, Lu YJ, Söhnlein P, Brousseau R, Laprade R, Masson L, Adang MJ. Ion channels formed in planar lipid bilayers by Bacillus thuringiensis toxins in the presence of Manduca sexta midgut receptors. FEBS Lett. 1997;412:270–6.CrossRefPubMedGoogle Scholar
  45. Siqueira HH, Nickerson KW, Moellenbeck D, Siegfried BD. Activity of gut proteinases from Cry1Ab-selected colonies of the European corn borer, Ostrinia nubilalis (Lepidoptera: Crambidae). Pest Manag Sci. 2004;60:1189–96.CrossRefPubMedGoogle Scholar
  46. Soberón M, Pardo-López L, López I, Gómez I, Tabashnik B, Bravo A. Engineering modified Bt toxins to counter insect resistance. Sciences. 2007;318:1640–2.CrossRefGoogle Scholar
  47. Soberón M, Gill SS, Bravo A. Signaling versus punching hole: how do Bacillus thuringiensis toxins kill insect midgut cells? Cell Mol Life Sci. 2009;66:1337–49.CrossRefPubMedGoogle Scholar
  48. Tabashnik BE, Huang F, Ghimire MN, Leonard BR, Siegfried BD, Randasamy M, Yang Y, Wu Y, Gahan L, Heckel DG, Bravo A, Soberón M. Efficacy of genetically modified Bt toxins against insects with different mechanism of resistance. Nat Biotechnol. 2011;29:1128–31.CrossRefPubMedGoogle Scholar
  49. Tabashnik BE, Brévault T, Carrière Y. Insect resistance to Bt crops: lessons from the first billion acres. Nat Biotechnol. 2013;31(6):510–21.CrossRefPubMedGoogle Scholar
  50. Tabashnik BE, Zhang M, Fabrick JA, Wu Y, Gao M, Huang F, Wei J, Zhang J, Yelich A, Unnithan GC, Bravo A, Soberón M, Carrière Y, Li X. Dual mode of action of Bt proteins: protoxin efficacy against resistant insects. Sci Rep. 2015;5:15107.CrossRefPubMedPubMedCentralGoogle Scholar
  51. Tay WT, Mahon RJ, Heckel DG, Walsh TK, Downes S, James WJ, Lee S-F, Reineke A, Williams AK, Gordon KHJ. Insect resistance to Bacillus thuringiensis toxin Cry2Ab is conferred by mutations in an ABC transporter subfamily A protein. PLoS Genet. 2015;11:e1005534.CrossRefPubMedPubMedCentralGoogle Scholar
  52. Vachon V, Prefontaine G, Coux F, Rang C, Marceau L, Masson L, Brousseau R, Frutos R, Schwartz JL, Laprade R. Role of helix 3 in pore formation by Bacillus thuringiensis insecticidal toxin Cry1Aa. Biochemistry. 2002;41:6178–84.CrossRefPubMedGoogle Scholar
  53. Xiao Y, Zhang T, Liu C, Heckel DG, Li X, Tabashnik BE, Wu K. Mis-splicing of the ABCC2 gene linked with Bt toxin resistance in Helicoverpa armigera. Sci Rep. 2014;4:6184.CrossRefPubMedPubMedCentralGoogle Scholar
  54. Xu X, Yu L, Wu Y. Disruption of a cadherin gene associated with resistance to Cry1Ac delta-endotoxin of Bacillus thuringiensis in Helicoverpa armigera. Appl Environ Microbiol. 2005;71:948–54.CrossRefPubMedPubMedCentralGoogle Scholar
  55. Yang YH, Yang YJ, Gao WY, Guo JJ, Wu YH, Wu YD. Introgression of a disrupted cadherin gene enables susceptible Helicoverpa armigera to obtain resistance to Bacillus thuringiensis toxin Cry1Ac. Bull Entomol Res. 2009;99:175–81.CrossRefPubMedGoogle Scholar
  56. Yang Y, Zhu YC, Ottea J, Husseneder C, Leonard BR, Abel C, Luttrell R, Huang F. Down regulation of a gene for cadherin, but not alkaline phosphatase, associated with Cry1Ab resistance in sugarcane borer Diatraea saccharalis. PLoS One. 2011;6:e25783.CrossRefPubMedPubMedCentralGoogle Scholar
  57. Zhang X, Candas M, Griko NB, Taussig R, Bulla Jr LA. A mechanism of cell death involving an adenylyl cyclase/PKA signaling pathway is induced by the Cry1Ab toxin of Bacillus thuringiensis. Proc Natl Acad Sci U S A. 2006;103:9897–902.CrossRefPubMedPubMedCentralGoogle Scholar
  58. Zhang S, Cheng H, Gao Y, Wang G, Liang G, Wu K. Mutation of an aminopeptidase N gene is associated with Helicoverpa armigera resistance to Bacillus thuringiensis Cry1Ac toxin. Insect Biochem Mol Biol. 2009;39:421–9.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2016

Authors and Affiliations

  • Mario Soberón
    • 1
  • Rose Monnerat
    • 2
  • Alejandra Bravo
    • 1
  1. 1.Instituto de BiotecnologiaUniversidad Nacional Autonoma de MéxicoCuernavacaMexico
  2. 2.Embrapa Recursos Genéticos e BiotecnologiaBrasília, DFBrazil

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