Localization of Bacillus thuringiensis Cry1A toxin-binding molecules in gypsy moth larval gut sections using fluorescence microscopy

Valaitis, Algimantas P.

doi:10.1016/j.jip.2011.07.001

Cited by 11 publications

(12 citation statements)

References 42 publications

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“…This result suggests that Cry4Ba does not rely on binding to cadherin for A. aegypti toxicity either because it relies on binding to another as yet unidentified midgut molecule for oligomer formation or because it can form functional oligomers just by protease activation. In the lepidopteran Lymantria dispar , Cry1Ac toxin does not bind to cadherin, but instead relies on the binding to a different midgut protein, an anionic glycoconjugate (BTR270) [26]. …”

Section: Discussionmentioning

confidence: 99%

Cadherin binding is not a limiting step for Bacillus thuringiensis subsp. israelensis Cry4Ba toxicity to Aedes aegypti larvae

Rodríguez-Almazán

Reyes

Zúñiga-Navarrete

et al. 2012

Biochemical Journal

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Bacillus thuringiensis subsp. israelensis produces three Cry toxins (Cry4Aa, Cry4Ba and Cry11Aa) that are active against Aedes aegypti larvae. The identification of the rate-limiting binding steps of Cry toxins that are used for insect control in the field, such as those of B. thuringiensis subsp. israelensis, should provide targets for improving insecticides against important insect pests. Previous studies showed that Cry11Aa binds to cadherin receptor fragment CR7–11 (cadherin repeats 7–11) with high affinity. Binding to cadherin has been proposed to facilitate Cry toxin oligomer formation. In the present study, we show that Cry4Ba binds to CR7–11 with 9-fold lower binding affinity compared with Cry11Aa. Oligomerization assays showed that Cry4Ba is capable of forming oligomers when proteolytically activated in vitro in the absence of the CR7–11 fragment in contrast with Cry11Aa that formed oligomers only in the presence of CR7–11. Pore-formation assays in planar lipid bilayers showed that Cry4Ba oligomers were proficient in opening ion channels. Finally, silencing the cadherin gene by dsRNA (double-stranded RNA) showed that silenced larvae were more tolerant to Cry11Aa in contrast with Cry4Ba, which showed similar toxic levels to those of control larvae. These findings show that cadherin binding is not a limiting step for Cry4Ba toxicity to A. aegypti larvae.

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Section: Discussionmentioning

confidence: 99%

Cadherin binding is not a limiting step for Bacillus thuringiensis subsp. israelensis Cry4Ba toxicity to Aedes aegypti larvae

Rodríguez-Almazán

Reyes

Zúñiga-Navarrete

et al. 2012

Biochemical Journal

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“…Members of the aminopeptidase N (APN) gene family of the insect order Lepidoptera (moths and butterflies) bind to insecticidal Cry toxins naturally produced by the bacterium Bacillus thuringiensis . Various studies on APN1 from Manduca sexta , Lymantria dispar , P. xylostella , H. armigera and Trichoplusia ni have consistently demonstrated that this protein is one of the midgut receptors for Cry1Ac. During our research on APN1 in H. armigera , 21 different linear transcripts were detected from two susceptible strains.…”

Section: Discussionmentioning

confidence: 98%

Newly identified APN splice isoforms suggest novel splicing mechanisms may underlie circRNA circularization in moth

et al. 2019

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Circular RNA (circ RNA ) have long been considered by‐products of splicing errors, but the coordination of RNA transcription and exon circularization events remains poorly understood. Here, we investigated this question using genes encoding aminopeptidases N ( APN s), which are receptors of Bacillus thuringiensis toxins, in the cotton bollworm, Helicoverpa armigera . We cloned and sequenced the cDNA of ten APN genes ( Ha APN 1‐10 ) located in the same APN gene cluster, and detected 20 and 14 novel splicing isoforms with exon skipping in Ha APN 1 and Ha APN 3 , respectively, whereas no or very few variants were found in the remaining genes. Further study identified 14 and 6 circular RNA (circ RNA ) in Ha APN 1 and Ha APN 3, respectively. Neither novel splicing isoforms nor circ RNA were detected in Ha APN 2 and Ha APN 5 . Distinct from the conventional GT / AG splicing signal, short co‐directional repeats were involved in the splicing of the linear and circular isoforms of Ha APN 1 and Ha APN 3 . Identification of the splice sites revealed that the linear isoforms may be related in some way to the circularization. Moreover, phylogenetic analysis and detection of circ RNA of the APN gene of the diamondback moth, Plutella xylostella ( Px APN 3 ), showed that circ RNA formation is relatively conserved during the lepidopteran evolutionary process. These results contribute to an improved understanding of lepidopteran APN s and this novel class of insect circ RNA .

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“…It has also been shown that ALP2 plays a dominant role in the susceptibility of S. exigua to the Cry2Aa toxin60. Among the several clusters of APNs present in lepidopteran insects, APN1 has been reported as an important functional receptor for Cry1Ac in several lepidopteran species424461, and APN3 and APN6 serve as functional candidate receptors for Cry1Ca in S. exigua 62. However, there is no direct or conflict evidence of the roles of other APNs in the mechanisms of Cry1A, Cry2A or other Cry toxins46.…”

Section: Discussionmentioning

confidence: 99%

Functional roles of cadherin, aminopeptidase-N and alkaline phosphatase from Helicoverpa armigera (Hübner) in the action mechanism of Bacillus thuringiensis Cry2Aa

Zhao

Yuan

Wei

et al. 2017

Sci Rep

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A pyramid strategy combining the Cry1A and Cry2A toxins in Bt crops has been widely used throughout the world to delay pest adaption to transgenic crops and broaden the insecticidal spectrum. Midgut membrane-bound cadherin (CAD), aminopeptidase-N (APN) and alkaline phosphatase (ALP) are important for Cry1A toxicity in some lepidopteran larvae, but the proteins that bind Cry2A in the midgut of target insects and their role in the Cry2A mechanism of action are still unclear. In this study, we found that heterologously expressed CAD, APN4 and ALP2 peptides from the midgut of Helicoverpa armigera could bind to the Cry2Aa toxin with a high affinity. Additionally, the efficiency of Cry2Aa insecticidal activity against H. armigera larvae was obviously reduced after the genes encoding these proteins were silenced with specific siRNAs: CAD- and ALP2-silenced larvae showed significantly similar reductions in mortality due to the Cry2Aa toxin (41.67% and 43.06%, respectively), whereas a larger reduction in mortality was observed in APN4-silenced larvae (61.11%) than in controls. These results suggest that CAD, APN4 and ALP2 are involved in the mechanism of action of Cry2Aa in H. armigera and may play important functional roles in the toxicity of the Cry2Aa toxin.

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Localization of Bacillus thuringiensis Cry1A toxin-binding molecules in gypsy moth larval gut sections using fluorescence microscopy

Cited by 11 publications

References 42 publications

Cadherin binding is not a limiting step for Bacillus thuringiensis subsp. israelensis Cry4Ba toxicity to Aedes aegypti larvae

Cadherin binding is not a limiting step for Bacillus thuringiensis subsp. israelensis Cry4Ba toxicity to Aedes aegypti larvae

Newly identified APN splice isoforms suggest novel splicing mechanisms may underlie circRNA circularization in moth

Functional roles of cadherin, aminopeptidase-N and alkaline phosphatase from Helicoverpa armigera (Hübner) in the action mechanism of Bacillus thuringiensis Cry2Aa

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