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Intermolecular Interaction Between Cry2Aa and Cyt1Aa and Its Effect on Larvicidal Activity Against Culex quinquefasciatus

  • Bideshi, Dennis K. (Department of Entomology, University of California) ;
  • Waldrop, Greer (Department of Biology, Undergraduate Program in Molecular, Cellular, and Developmental Biology, University of Louisville) ;
  • Fernandez-Luna, Maria Teresa (Department of Entomology, University of California) ;
  • Diaz-Mendoza, Mercedes (Department of Entomology, University of California) ;
  • Wirth, Margaret C. (Department of Entomology, University of California) ;
  • Johnson, Jeffrey J. (Department of Entomology, University of California) ;
  • Park, Hyun-Woo (Department of Entomology, University of California) ;
  • Federici, Brian A. (Department of Entomology, University of California)
  • Received : 2013.01.23
  • Accepted : 2013.05.05
  • Published : 2013.08.28

Abstract

The Cyt1Aa protein of Bacillus thuringiensis susbp. israelensis elaborates demonstrable toxicity to mosquito larvae, but more importantly, it enhances the larvicidal activity of this species Cry proteins (Cry11Aa, Cry4Aa, and Cry4Ba) and delays the phenotypic expression of resistance to these that has evolved in Culex quinquefasciatus. It is also known that Cyt1Aa, which is highly lipophilic, synergizes Cry11Aa by functioning as a surrogate membrane-bound receptor for the latter protein. Little is known, however, about whether Cyt1Aa can interact similarly with other Cry proteins not primarily mosquitocidal; for example, Cry2Aa, which is active against lepidopteran larvae, but essentially inactive or has very low toxicity to mosquito larvae. Here we demonstrate by ligand binding and enzyme-linked immunosorbent assays that Cyt1Aa and Cry2Aa form intermolecular complexes in vitro, and in addition show that Cyt1Aa facilitates binding of Cry2Aa throughout the midgut of C. quinquefasciatus larvae. As Cry2Aa and Cry11Aa share structural similarity in domain II, the interaction between Cyt1Aa and Cry2Aa could be a result of a similar mechanism previously proposed for Cry11Aa and Cyt1Aa. Finally, despite the observed interaction between Cry2Aa and Cyt1Aa, only a 2-fold enhancement in toxicity resulted against C. quinquefasciatus. Regardless, our results suggest that Cry2Aa could be a useful component of mosquitocidal endotoxin complements being developed for recombinant strains of B. thuringiensis subsp. israelensis and B. sphaericus aimed at improving the efficacy of commercial products and avoiding resistance.

Keywords

References

  1. Bradford MM. 1976. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem. 72: 248-254. https://doi.org/10.1016/0003-2697(76)90527-3
  2. Bravo A, Gill SS, Soberon M. 2007. Mode of action of Bacillus thuringiensis Cry and Cyt toxins and their potential for insect control. Toxicon. 15: 423-435.
  3. Butko P. 2003. Cytolytic toxin Cyt1A and its mechanism of membrane damage: data and hypothesis. Appl. Environ. Microbiol. 69: 2415-2422. https://doi.org/10.1128/AEM.69.5.2415-2422.2003
  4. Crickmore N, Bone EJ, Williams JA, Ellar DJ. 1995. Contribution of the individual components of the $\delta$-endotoxin crystal to the mosquitocidal activity of Bacillus thuringiensis. FEMS Microbiol. Lett. 131: 249-254.
  5. Diaz-Mendoza M, Bideshi DK, Federici BA. 2012. A 54-kDa protein encoded by pBtoxis is required for parasporal body structural integrity in Bacillus thuringiensis subsp. israelensis. J. Bacteriol. 194: 1562-1571. https://doi.org/10.1128/JB.06095-11
  6. Federici BA, Park HW, Bideshi DK, Wirth MC, Johnson JJ, Sakano Y, et al. 2007. Developing recombinant bacteria for control of mosquito larvae. J. Am. Mosq. Control Assoc. 23: 164-175. https://doi.org/10.2987/8756-971X(2007)23[164:DRBFCO]2.0.CO;2
  7. Federici BA, Park HW, Sakano Y. 2006. Insecticidal protein crystals of Bacillus thuringiensis, pp. 195-236. In Shively JM (ed.). Microbiology Monographs Inclusions in Prokaryotes. Springer-Verlag, Berlin, Heidelberg.
  8. Fernandez LE, Perez C, Segovia L, Rodriguez MH, Gill SS, Bravo A, et al. 2005. Cry11Aa toxin from Bacillus thuringiensis binds its receptor in Aedes aegypti mosquito larvae through loop a-8 of domain II. FEBS Lett. 579: 3508-3514. https://doi.org/10.1016/j.febslet.2005.05.032
  9. Finney D. 1971. Probit Analysis. Cambridge University Press, Cambridge, England.
  10. Ge B, Bideshi D, Moar WJ, Federici BA. 1998. Differential effects of helper proteins encoded by the cry2A and cry11A operons on the formation of Cry2A inclusions in Bacillus thuringiensis. FEMS Microbiol. Lett. 165: 35-41. https://doi.org/10.1111/j.1574-6968.1998.tb13124.x
  11. Georghiou GP, Wirth MC. 1997. Influence of exposure to single versus multiple toxins of Bacillus thuringiensis subsp. israelensis on development of resistance in the mosquito Culex quinquefasciatus (Diptera: Culicidae). Appl. Environ. Microbiol. 63: 1095-1101.
  12. Guerchicoff A, Delecluse A, Rubinstein CP. 2001. The Bacillus thuringiensis cyt genes for hemolytic endotoxin constitute a gene family. Appl. Environ. Microbiol. 67: 1090-1096. https://doi.org/10.1128/AEM.67.3.1090-1096.2001
  13. Ibarra J, Federici BA. 1986. Isolation of a relatively nontoxic 65-kilodalton protein inclusion from the parasporal body of Bacillus thuringiensis subsp. israelensis. J. Bacteriol. 165: 527-533. https://doi.org/10.1128/jb.165.2.527-533.1986
  14. Knowles BH, Blatt MR, Tester M, Horsnell JM, Caroll J, Menestrina G, et al. 1992. A cytolytic delta-endotoxin from Bacillus thuringiensis var. israelensis forms cation-selective channels in planar lipid bilayers. FEBS Lett. 244: 259-262.
  15. Laemmli UK. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680-685. https://doi.org/10.1038/227680a0
  16. Lereclus A, Arantes O, Chaufaux J, Lecadet MM. 1989. Transformation and expression of a cloned d-endotoxin gene in Bacillus thuringiensis. FEMS Microbiol. Lett. 60: 211-218.
  17. Li K, Pandelakis AK, Ellar DJ. 1996. Structure of the mosquitocidal d-endotoxin CytB from Bacillus thuringiensis sp. kyushuensis and implications for membrane pore formation. J. Mol. Biol. 257: 129-152. https://doi.org/10.1006/jmbi.1996.0152
  18. Li X, Nevels KJ, Gryczynski Z, Gryczynski I, Pusztai-Carey M, Xie D, et al. 2009. A molecular dynamic model of the Bt toxin Cyt1A and its validation by resonance energy transfer. Biophys. Chem. 144: 53-61. https://doi.org/10.1016/j.bpc.2009.06.005
  19. Manceva SD, Pusztai-Carey M, Russo PS, Butko P. 2005. A detergent-like mechanism of action of the cytolytic toxin Cyt1A from Bacillus thuringiensis var. israelensis. Biochemistry 44: 589-597. https://doi.org/10.1021/bi048493y
  20. Meyer SK, Tabashnik BE, Liu YB, Wirth MC, Federici BA. 2001. Cyt1A from Bacillus thuringiensis lacks toxicity to susceptible and resistant larvae of diamondback moth (Plutella xylostella) and pink bollworm (Pectinophora gossypiella). Appl. Environ. Microbiol. 67: 462-463. https://doi.org/10.1128/AEM.67.1.462-463.2001
  21. Moar WJ, Trumble JT, Hice RH, Backman PA. 1994. Insecticidal activity of the CryIIA protein form the NRD-12 isolates of Bacillus thuringiensis subsp. kurstaki expressed in Escherichia coli and Bacillus thuringiensis and in a leafcolonizing strain of Bacillus cereus. Appl. Environ. Microbiol. 3: 896-902.
  22. Morse RJ, Yamamoto T, Stroud RM. 2001. Structure of Cry2Aa suggests an unexpected receptor binding epitope. Structure 9: 409-417. https://doi.org/10.1016/S0969-2126(01)00601-3
  23. Park HW, Bideshi DK, Federici BA. 2003. Recombinant strain of Bacillus thuringiensis producing Cyt1A, Cry11B and the Bacillus sphaericus toxin. Appl. Environ. Microbiol. 69: 1331-1334. https://doi.org/10.1128/AEM.69.2.1331-1334.2003
  24. Park HW, Bideshi DK, Wirth MC, Johnson JJ, Walton WE, Federici BA. 2005. Recombinant larvicidal bacteria with markedly improved efficacy against Culex vectors of West Nile virus. Am. J. Trop. Med. Hyg. 72: 732-738.
  25. Perez C, Fernandez LE, Sun L, Folch LJ, Gill SS, Soberon M, et al. 2005. Bacillus thuringiensis subsp. israelensis Cyt1Aa synergizes Cry11A toxin by functioning as a membranebound receptor. Proc. Natl. Acad. Sci. USA 102: 18303-18308. https://doi.org/10.1073/pnas.0505494102
  26. Poncet S, Delecluse A, Klier A, Rapoport G. 1994. Evaluation of synergistic interactions among CryIVA, CryIVB, and CryIVD toxic components of Bacillus thuringiensis subsp. israelensis crystals. J. Invertebr. Pathol. 66: 131-135.
  27. Roh JY, Choi JY, Li MS, Jin BR, Je JH. 2007. Bacillus thuringiensis as a specific, safe, and effective tool for insect pest control. J. Microbiol. Biotechnol. 17: 547-549.
  28. Schnepf E, Crickmore N, van Rie J, Lereclus D, Baum J, Feitelson J, et al. 1998. Bacillus thuringiensis and its pesticidal crystal proteins. Microbiol. Mol. Biol. Rev. 62: 775-806.
  29. Soberon M, Gill SS, Bravo A. 2009. Signaling versus punching holes: how do Bacillus thuringiensis toxins kill insect midgut cells? Cell. Mol. Life Sci. 66: 1337-1349. https://doi.org/10.1007/s00018-008-8330-9
  30. Swiecicka I. 2008. Natural occurrence of Bacillus thuringiensis and Bacillus cereus in eukaryotic organisms: a case of symbiosis. Biocontrol Sci. Technol. 18: 221-239. https://doi.org/10.1080/09583150801942334
  31. Tabashnik BE. 1992. Evaluation of synergism among Bacillus thuringiensis toxins. Appl. Environ. Microbiol. 58: 3343-3346.
  32. Thomas WL, Ellar DJ. 1983. Mechanism of action of Bacillus thuringiensis var. israelensis insecticidal delta-endotoxins. FEBS Lett. 154: 362-368. https://doi.org/10.1016/0014-5793(83)80183-5
  33. Widner WR, Whiteley HR. 1989. Two highly related insecticidal crystal proteins of Bacillus thuringiensis subsp. kurstaki possess different host range specificities. J. Bacteriol. 171: 965-974. https://doi.org/10.1128/jb.171.2.965-974.1989
  34. Widner WR, Whiteley HR. 1990. Location of the dipteran specificity region in a lepidopteran-dipteran crystal protein from Bacillus thuringiensis. J. Bacteriol. 172: 2826-2832. https://doi.org/10.1128/jb.172.6.2826-2832.1990
  35. Wirth MC, Delecluse A, Walton WE. 2004. Laboratory selection for resistance to Bacillus thuringiensis subsp. jegathesan or a component toxin, Cry11B, in Culex quinquefasciatus (Diptera: Culicidae). J. Med. Entomol. 41: 435-441. https://doi.org/10.1603/0022-2585-41.3.435
  36. Wirth MC, Geroghiou GP, Federici BA. 1997. Cyt1A enables CryIVD endotoxins of Bacillus thuringiensis to overcome high levels of CryIV resistance in the mosquito Culex quinquefasciatus. Proc. Natl. Acad. Sci. USA 94: 10536-10540. https://doi.org/10.1073/pnas.94.20.10536
  37. Wirth MC, Park HW, Walton WE, Federici BA. 2005. Cyt1A of Bacillus thuringiensis delays evolution of resistance to Cry11A in the mosquito Culex quinquefasciatus. Appl. Environ. Microbiol. 71: 185-189. https://doi.org/10.1128/AEM.71.1.185-189.2005
  38. Wirth MC, Jiannino JA, Federici BA, Walton WE. 2004. Synergy between toxins of Bacillus thuringiensis subsp. israelensis and Bacillus sphaericus. J. Med. Entomol. 41: 935-941. https://doi.org/10.1603/0022-2585-41.5.935
  39. Wu D, Federici BA. 1993. The 20-kilodalton protein preserves cell viability and promotes CytA crystal formation during sporulation in Bacillus thuringiensis. J. Bacteriol. 175: 5276-5280. https://doi.org/10.1128/jb.175.16.5276-5280.1993
  40. Zghal R Z, T ounsi S, J aoua J . 2006. C haracterization o f a Cry4Ba-type gene of Bacillus thuringiensis israelensis and evidence of the synergistic larvicidal activity of its encoded protein Cry2A d-endotoxin of B. thuringiensis kurstaki on Culex pipiens (common house mosquito). Biotechnol. Appl. Biochem. 44: 19-25. https://doi.org/10.1042/BA20050134

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