Bacillus thuringiensis subsp. israelensis (Bti) has been used in the field for over twenty years without resistance development in any target insect. In contrast, mosquito resistance to B. sphaericus has been observed in the field in many countries. This remarkable difference in the propensity to develop resistance to two mosquitocidal Bacillus strains is likely due to the presence of multiple toxins in Bti. However, mosquitoes are able to rapidly develop resistance to individual toxins from this strain. A major reason for the apparent inability of mosquitoes to develop resistance to Bti is the presence of cytolytic (Cyt) toxins in this and other mosquitocidal strains. During the last grant period we showed that key to the lack of mosquito resistance to Bti was how two toxins in this strain, namely Cry11A and Cyt1A toxins interact. We showed Cyt1A acts as a surrogate receptor for Cry11Aa. Importantly Cyt1Aa binds through Cry11A loop domains that are also involved in binding endogenous Aedes aegypti receptor proteins. Our research showed Aedes receptor proteins include cadherin, alkaline phosphatases (ALP), and aminopeptidases (APNs). All three classes bind mosquitocidal toxins with relatively high affinity, unlike in lepidopterans where only cadherin binds with high affinity. This implies both APN and ALP of mosquitoes could act as primary rather than as secondary receptors as in moths, suggesting a possible difference in the mode of action of mosquitocidal and lepidopteran Cry toxins. We believe however, there is conservation in the mode of action of Bt Cry toxins. Therefore we hypothesize a similar mode of action of action occurs in mosquitoes as in lepidopterans. In this proposal we plan to test this hypothesis. Consequently, we hypothesize that: (i) cadherin is a key protein which mediates initial binding to mosquitocidal Cry toxins and is essential for larval toxicity;ii) ALPs (and APNs) act as secondary receptors that allow toxin targeting to the cell membrane. Also in the previous proposal we showed Cyt1A plays a critical role in acting as a surrogate receptor for Cry11Aa. We therefore hypothesize that (iii) Cyt1A also is a receptor for other Cry toxins in Bti, and synergizes the toxicity of mosquitocidal Cry toxins. We will test this hypothesis and also elucidate the mechanism by which Cyt toxins insert into the membrane to act as surrogate receptors. This project is also a proposal to continue a successful collaboration between three different investigators to best use the expertise of each laboratory.

Public Health Relevance

B. thuringiensis subsp. israelensis are used for control of human disease vectors, such as species of Simulium, Aedes, Culex, and Anopheles. Elucidating mechanisms of Cry toxin action in aids our understanding of how mosquito and black fly control is achieved and also enables us to determine mechanisms by which resistance can occur. This mode of Cyt1A toxin action investigation also illuminates mechanism of synergism between mosquitocidal Cry and Cyt toxins. Finally, elucidation of mechanisms by which the Cyt1Aa toxin acts also will help us understand how other cytolytic toxins of human health significance cause toxicity.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
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Vector Biology Study Section (VB)
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Costero, Adriana
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University of California Riverside
Anatomy/Cell Biology
Schools of Earth Sciences/Natur
United States
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Pacheco, Sabino; Gómez, Isabel; Sánchez, Jorge et al. (2017) An Intramolecular Salt Bridge in Bacillus thuringiensis Cry4Ba Toxin Is Involved in the Stability of Helix ?-3, Which Is Needed for Oligomerization and Insecticidal Activity. Appl Environ Microbiol 83:
Chen, Jianwu; Aimanova, Karly; Gill, Sarjeet S (2017) Functional characterization of Aedes aegypti alkaline phosphatase ALP1 involved in the toxicity of Cry toxins from Bacillus thuringiensis subsp. israelensis and jegathesan. Peptides 98:78-85
Canton, Pablo Emiliano; Cancino-Rodezno, Angeles; Gill, Sarjeet S et al. (2015) Transcriptional cellular responses in midgut tissue of Aedes aegypti larvae following intoxication with Cry11Aa toxin from Bacillus thuringiensis. BMC Genomics 16:1042
Lee, Su-Bum; Chen, Jianwu; Aimanova, Karlygash G et al. (2015) Aedes cadherin mediates the in vivo toxicity of the Cry11Aa toxin to Aedes aegypti. Peptides 68:140-147
Lee, Su-Bum; Aimanova, Karlygash G; Gill, Sarjeet S (2014) Alkaline phosphatases and aminopeptidases are altered in a Cry11Aa resistant strain of Aedes aegypti. Insect Biochem Mol Biol 54:112-21
Qureshi, Nadia; Chawla, Swati; Likitvivatanavong, Supaporn et al. (2014) The cry toxin operon of Clostridium bifermentans subsp. malaysia is highly toxic to Aedes Larval Mosquitoes. Appl Environ Microbiol 80:5689-97
Gómez, Isabel; Sánchez, Jorge; Muñoz-Garay, Carlos et al. (2014) Bacillus thuringiensis Cry1A toxins are versatile proteins with multiple modes of action: two distinct pre-pores are involved in toxicity. Biochem J 459:383-96
Cantón, Pablo Emiliano; López-Díaz, Jazmin A; Gill, Sarjeet S et al. (2014) Membrane binding and oligomer membrane insertion are necessary but insufficient for Bacillus thuringiensis Cyt1Aa toxicity. Peptides 53:286-91
Bedoya-Pérez, Leidy P; Cancino-Rodezno, Angeles; Flores-Escobar, Biviana et al. (2013) Role of UPR pathway in defense response of Aedes aegypti against Cry11Aa toxin from Bacillus thuringiensis. Int J Mol Sci 14:8467-78
López-Diaz, Jazmin A; Cantón, Pablo Emiliano; Gill, Sarjeet S et al. (2013) Oligomerization is a key step in Cyt1Aa membrane insertion and toxicity but not necessary to synergize Cry11Aa toxicity in Aedes aegypti larvae. Environ Microbiol 15:3030-9

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