Role of Bacillus thuringiensis Cry1A toxins domains in the binding to the ABCC2 receptor from Spodoptera exigua

Martínez-Solís, María; Pinos, Daniel; Endo, Haruka; Portugal, Leivi; Satō, Ryōichi; Ferré, Juan; Herrero, Salvador; Hernández‐Martínez, Patricia

doi:10.1016/j.ibmb.2018.07.006

Cited by 23 publications

(34 citation statements)

References 63 publications

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“…The toxicity data obtained were expected based on the published data [29,44,49,50]. Moreover, Sf21 cells were also used in this study, on the bases that Cry1Ia and Cry1Ab are not toxic for them [40,41].…”

Section: Discussionmentioning

confidence: 99%

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Study of the Bacillus thuringiensis Cry1Ia Protein Oligomerization Promoted by Midgut Brush Border Membrane Vesicles of Lepidopteran and Coleopteran Insects, or Cultured Insect Cells

Khorramnejad

Domínguez-Arrizabalaga

Caballero

et al. 2020

Toxins

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Bacillus thuringiensis (Bt) produces insecticidal proteins that are either secreted during the vegetative growth phase or accumulated in the crystal inclusions (Cry proteins) in the stationary phase. Cry1I proteins share the three domain (3D) structure typical of crystal proteins but are secreted to the media early in the stationary growth phase. In the generally accepted mode of action of 3D Cry proteins (sequential binding model), the formation of an oligomer (tetramer) has been described as a major step, necessary for pore formation and subsequent toxicity. To know if this could be extended to Cry1I proteins, the formation of Cry1Ia oligomers was studied by Western blot, after the incubation of trypsin activated Cry1Ia with insect brush border membrane vesicles (BBMV) or insect cultured cells, using Cry1Ab as control. Our results showed that Cry1Ia oligomers were observed only after incubation with susceptible coleopteran BBMV, but not following incubation with susceptible lepidopteran BBMV or non-susceptible Sf21 insect cells, while Cry1Ab oligomers were persistently detected after incubation with all insect tissues tested, regardless of its host susceptibility. The data suggested oligomerization may not necessarily be a requirement for the toxicity of Cry1I proteins.

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Section: Discussionmentioning

confidence: 99%

“…It has been described that Sf21 cells (insect cultured cells derived from S. frugiperda ovaries) are tolerant to both Cry1Ab and Cry1Ia proteins [40,41]. These insect cells were selected in order to check if oligomer formation could be promoted by tolerant insect tissues.…”

Section: Oligomerization Promoted By Sf21 Insect Cellsmentioning

confidence: 99%

Study of the Bacillus thuringiensis Cry1Ia Protein Oligomerization Promoted by Midgut Brush Border Membrane Vesicles of Lepidopteran and Coleopteran Insects, or Cultured Insect Cells

Khorramnejad

Domínguez-Arrizabalaga

Caballero

et al. 2020

Toxins

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“…Absorbance was measured at 490 nm (Infinite m200, Tecan, Mannedorf, Switzerland). The percentage of viable cells was obtained as described elsewhere [ 48 ]. Mean values in the transfected cells against the non-transfected cells were compared by Student’s t -test at 5% level of significance.…”

Section: Methodsmentioning

confidence: 99%

Reduced Membrane-Bound Alkaline Phosphatase Does Not Affect Binding of Vip3Aa in a Heliothis virescens Resistant Colony

Pinos

Chakroun

Millán-Leiva

et al. 2020

Toxins

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The Vip3Aa insecticidal protein from Bacillus thuringiensis (Bt) is produced by specific transgenic corn and cotton varieties for efficient control of target lepidopteran pests. The main threat to this technology is the evolution of resistance in targeted insect pests and understanding the mechanistic basis of resistance is crucial to deploy the most appropriate strategies for resistance management. In this work, we tested whether alteration of membrane receptors in the insect midgut might explain the >2000-fold Vip3Aa resistance phenotype in a laboratory-selected colony of Heliothis virescens (Vip-Sel). Binding of 125I-labeled Vip3Aa to brush border membrane vesicles (BBMV) from 3rd instar larvae from Vip-Sel was not significantly different from binding in the reference susceptible colony. Interestingly, BBMV from Vip-Sel larvae showed dramatically reduced levels of membrane-bound alkaline phosphatase (mALP) activity, which was further confirmed by a strong downregulation of the membrane-bound alkaline phosphatase 1 (HvmALP1) gene. However, the involvement of HvmALP1 as a receptor for the Vip3Aa protein was not supported by results from ligand blotting and viability assays with insect cells expressing HvmALP1.

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“…Mutations in ABC transporter subfamily C2 (ABCC2) are associated with Bt (Cry1A toxins) resistance in a number of lepidopteran species [11][12][13][14][15][16][17]. The receptor functionality of ABCC2 for Cry1A toxins has been well characterized using heterologous expression in cultured cell lines, manipulating mRNA abundance with RNA interference and with transgenic Drosophila [14,[18][19][20][21][22]. ABC transporter subfamily C3 (ABCC3), a paralog of ABCC2, has also been reported to mediate Cry1A toxicity [14,[22][23][24].…”

Section: Introductionmentioning

confidence: 99%

Resistance to Bacillus thuringiensis Cry1Ac toxin requires mutations in two Plutella xylostella ATP-binding cassette transporter paralogs

Xiang

et al. 2020

PLoS Pathog

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The diamondback moth, Plutella xylostella, is a cosmopolitan pest and the first species to develop field resistance to toxins from the gram-positive bacterium Bacillus thuringiensis (Bt). Although previous work has suggested that mutations of ATP-binding cassette transporter subfamily C2 (ABCC2) or C3 (ABCC3) genes can confer Cry1Ac resistance, here we reveal that P. xylostella requires combined mutations in both PxABCC2 and PxABCC3 to achieve high-level Cry1Ac resistance, rather than simply a mutation of either gene. We identified natural mutations of PxABCC2 and PxABCC3 that concurrently occurred in a Cry1Acresistant strain (Cry1S1000) of P. xylostella, with a mutation (R A2) causing the mis-splicing of PxABCC2 and another mutation (R A3) leading to the premature termination of PxABCC3. Genetic linkage analysis showed that R A2 and R A3 were tightly linked to Cry1Ac resistance. Introgression of R A2 and R A3 enabled a susceptible strain (G88) of P. xylostella to obtain high resistance to Cry1Ac, confirming that these genes confer resistance. To further support the role of PxABCC2 and PxABCC3 in Cry1Ac resistance, frameshift mutations were introduced into PxABCC2 and PxABCC3 singly and in combination in the G88 strain with CRISPR/Cas9 mediated mutagenesis. Bioassays of CRISPR-based mutant strains, plus genetic complementation tests, demonstrated that the deletion of PxABCC2 or PxABCC3 alone provided < 4-fold tolerance to Cry1Ac, while disruption of both genes together conferred >8,000-fold resistance to Cry1Ac, suggesting the redundant/complementary roles of PxABCC2 and PxABCC3. This work advances our understanding of Bt resistance in P. xylostella by demonstrating mutations within both PxABCC2 and PxABCC3 genes are required for high-level Cry1Ac resistance.

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Role of Bacillus thuringiensis Cry1A toxins domains in the binding to the ABCC2 receptor from Spodoptera exigua

Cited by 23 publications

References 63 publications

Study of the Bacillus thuringiensis Cry1Ia Protein Oligomerization Promoted by Midgut Brush Border Membrane Vesicles of Lepidopteran and Coleopteran Insects, or Cultured Insect Cells

Study of the Bacillus thuringiensis Cry1Ia Protein Oligomerization Promoted by Midgut Brush Border Membrane Vesicles of Lepidopteran and Coleopteran Insects, or Cultured Insect Cells

Reduced Membrane-Bound Alkaline Phosphatase Does Not Affect Binding of Vip3Aa in a Heliothis virescens Resistant Colony

Resistance to Bacillus thuringiensis Cry1Ac toxin requires mutations in two Plutella xylostella ATP-binding cassette transporter paralogs

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