scholarly journals Toxicity and Mode of Action of Bacillus thuringiensis Cry Proteins in the Mediterranean Corn Borer, Sesamia nonagrioides (Lefebvre)

2006 ◽  
Vol 72 (4) ◽  
pp. 2594-2600 ◽  
Author(s):  
Joel González-Cabrera ◽  
Gema P. Farinós ◽  
Silvia Caccia ◽  
Mercedes Díaz-Mendoza ◽  
Pedro Castañera ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Binding Site ◽  
Mode Of Action ◽  
Membrane Vesicles ◽  
Bt Corn ◽  
Sesamia Nonagrioides ◽  
Cry Protein ◽  
Cry Proteins ◽  
Corn Borer ◽  
Mediterranean Countries

ABSTRACT Sesamia nonagrioides is one of the most damaging pests of corn in Spain and other Mediterranean countries. Bt corn expressing the Bacillus thuringiensis Cry1Ab toxin is being grown on about 58,000 ha in Spain. Here we studied the mode of action of this Cry protein on S. nonagrioides (binding to specific receptors, stability of binding, and pore formation) and the modes of action of other Cry proteins that were found to be active in this work (Cry1Ac, Cry1Ca, and Cry1Fa). Binding assays were performed with 125I- or biotin-labeled toxins and larval brush border membrane vesicles (BBMV). Competition experiments indicated that these toxins bind specifically and that Cry1Aa, Cry1Ab, and Cry1Ac share a binding site. Cry1Ca and Cry1Fa bind to different sites. In addition, Cry1Fa binds to Cry1A's binding site with very low affinity and vice versa. Binding of Cry1Ab and Cry1Ac was found to be stable over time, which indicates that the observed binding is irreversible. The pore-forming activity of Cry proteins on BBMV was determined using the voltage-sensitive fluorescent dye DiSC3(5). Membrane permeability increased in the presence of the active toxins Cry1Ab and Cry1Fa but not in the presence of the nonactive toxin Cry1Da. In terms of resistance management, based on our results and the fact that Cry1Ca is not toxic to Ostrinia nubilalis, we recommend pyramiding of Cry1Ab with Cry1Fa in the same Bt corn plant for better long-term control of corn borers.

scholarly journals Rearrangement of N-Terminal α-Helices of Bacillus thuringiensis Cry1Ab Toxin Essential for Oligomer Assembly and Toxicity

Toxins ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 647
Author(s):  
Sabino Pacheco ◽  
Jean Piere Jesus Quiliche ◽  
Isabel Gómez ◽  
Jorge Sánchez ◽  
Mario Soberón ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Conformational Changes ◽  
Membrane Integrity ◽  
Resonance Energy Transfer ◽  
Resonance Energy ◽  
Membrane Vesicles ◽  
Insect Midgut ◽  
Cry Proteins ◽  
Cry1ab Toxin ◽  
Domain I

Cry proteins produced by Bacillus thuringiensis are pore-forming toxins that disrupt the membrane integrity of insect midgut cells. The structure of such pore is unknown, but it has been shown that domain I is responsible for oligomerization, membrane insertion and pore formation activity. Specifically, it was proposed that some N-terminal α-helices are lost, leading to conformational changes that trigger oligomerization. We designed a series of mutants to further analyze the molecular rearrangements at the N-terminal region of Cry1Ab toxin that lead to oligomer assembly. For this purpose, we introduced Cys residues at specific positions within α-helices of domain I for their specific labeling with extrinsic fluorophores to perform Föster resonance energy transfer analysis to fluorescent labeled Lys residues located in Domains II–III, or for disulfide bridges formation to restrict mobility of conformational changes. Our data support that helix α-1 of domain I is cleaved out and swings away from the toxin core upon binding with Manduca sexta brush border membrane vesicles. That movement of helix α-2b is also required for the conformational changes involved in oligomerization. These observations are consistent with a model proposing that helices α-2b and α-3 form an extended helix α-3 necessary for oligomer assembly of Cry toxins.

Toxicity of Transgenic Corn Tissues Expressing Insecticidal Protein of Bacillus thuringiensis Berliner to Southwestern Corn Borer (Lepidoptera: Crambidae) in the Laboratory

2005 ◽  
Vol 40 (3) ◽  
pp. 307-315 ◽  
Author(s):  
Kerry C. Allen ◽  
Henry N. Pitre
Keyword(s):  
Bacillus Thuringiensis ◽  
Tissue Type ◽  
Bt Corn ◽  
Transgenic Corn ◽  
Laboratory Studies ◽  
Negative Effects ◽  
Diatraea Grandiosella ◽  
Corn Borer ◽  
Bt Toxin ◽  
Southwestern Corn Borer

Laboratory studies were conducted to compare the response of Diatraea grandiosella (Dyar) larvae to lyophilized transgenic corn tissue expressing the Cry1Ab endotoxin protein of Bacillus thuringiensis (Berliner) (Bt) with conventional corn tissue incorporated into a BioServ™ artificial diet. Whorl leaf, stalk, shank, husk, silk or kernel tissues were tested independently in the diet. Larvae fed diet containing conventional corn weighed more and were longer in length than larvae fed diet containing Bt corn for all tissue types included in the study. The number of larvae that survived depended on the tissue type and age of the tissue. Larvae fed diet with kernel tissue expressing Bt toxin had greater weight and body length than larvae fed the other Bt tissue types. The negative effects of Bt corn tissues expressing the Cry1Ab endotoxin protein on growth of D. grandiosella was observed, even at the diluted concentrations of toxic tissues incorporated into the diet in this study.

scholarly journals Extent of Variation of the Bacillus thuringiensis Toxin Reservoir: the Case of the Geranium Bronze, Cacyreus marshalli Butler (Lepidoptera: Lycaenidae)

2002 ◽  
Vol 68 (8) ◽  
pp. 4090-4094 ◽  
Author(s):  
Salvador Herrero ◽  
Marisé Borja ◽  
Juan Ferré
Keyword(s):  
Bacillus Thuringiensis ◽  
Binding Site ◽  
Management Strategies ◽  
Resistance Management ◽  
Cross Resistance ◽  
Binding Studies ◽  
Cry Proteins ◽  
Combined Use ◽  
The Common

ABSTRACT Despite the fact that around 200 cry genes from Bacillus thuringiensis have already been cloned, only a few Cry proteins are toxic towards a given pest. A crucial step in the mode of action of Cry proteins is binding to specific sites in the midgut of susceptible insects. Binding studies in insects that have developed cross-resistance discourage the combined use of Cry proteins sharing the same binding site. If resistance management strategies are to be implemented, the arsenal of Cry proteins suitable to control a given pest may be not so vast as it might seem at first. The present study evaluates the potential of B. thuringiensis for the control of a new pest, the geranium bronze (Cacyreus marshalli Butler), a butterfly that is threatening the popularity of geraniums in Spain. Eleven of the most common Cry proteins from the three lepidopteran-active Cry families (Cry1, Cry2, and Cry9) were tested against the geranium bronze for their toxicity and binding site relationships. Using 125I-labeled Cry1A proteins we found that, of the seven most active Cry proteins, six competed for binding to the same site. For the long-term control of the geranium bronze with B. thuringiensis-based insecticides it would be advisable to combine any of the Cry proteins sharing the binding site (preferably Cry1Ab, since it is the most toxic) with those not competing for the same site. Cry1Ba would be the best choice of these proteins, since it is significantly more toxic than the others not binding to the common site.

Influence of Transgenic Corn Expressing Insecticidal Protein of Bacillus thuringiensis Berliner on Natural Populations of Corn Earworm (Lepidoptera: Noctuidae) and Southwestern Corn Borer (Lepidoptera: Crambidae)

2006 ◽  
Vol 41 (3) ◽  
pp. 221-231 ◽  
Author(s):  
Kerry C. Allen ◽  
Henry N. Pitre
Keyword(s):  
Bacillus Thuringiensis ◽  
Helicoverpa Zea ◽  
Larval Growth ◽  
Natural Populations ◽  
Corn Earworm ◽  
Bt Corn ◽  
Transgenic Corn ◽  
Diatraea Grandiosella ◽  
Corn Hybrids ◽  
Corn Borer

A 2-yr study was conducted to measure the influence of transgenic corn, Zea mays L., expressing the CrylAb endotoxin of Bacillus thuringiensis (Berliner) (Bt) by means of Event MON810 on natural populations of Helicoverpa zea (Boddie) and Diatraea grandiosella (Dyar). The studies were conducted at Leland and Morgan City, MS, in 1999 and at Morgan City in 2000. Although total numbers of H. zea larvae were not significantly different on transgenic corn hybrids compared with their near-isogenic parent lines, fewer large larvae were found on the transgenic hybrids. Differences in H. zea larval growth were noticeable when larvae fed on Bt corn vs non-Bt corn. The delay in larval growth for insects within a single generation, which could possibly result in asynchronous mating between insecticide resistant and susceptible insects, was observed for larvae feeding on plants expressing the Bt toxin. Diatraea grandiosella caused limited damage to the transgenic corn hybrids compared with their near-isogenic parent lines. Yields were not significantly greater for the Bt corn hybrids compared with their near-isogenic parent lines. Yields were not significantly greater for the Bt corn hybrids compared with the near-isogenic, non-Bt corn parents; however, there was a trend toward higher yields for Bt hybrids compared with their near-isogenic non-Bt parents.

scholarly journals Binding Analyses of Bacillus thuringiensis Cry δ-Endotoxins Using Brush Border Membrane Vesicles of Ostrinia nubilalis

2001 ◽  
Vol 67 (2) ◽  
pp. 872-879 ◽  
Author(s):  
Gang Hua ◽  
Luke Masson ◽  
Juan Luis Jurat-Fuentes ◽  
George Schwab ◽  
Michael J. Adang
Keyword(s):  
Bacillus Thuringiensis ◽  
Binding Site ◽  
Brush Border ◽  
Ostrinia Nubilalis ◽  
European Corn Borer ◽  
Binding Proteins ◽  
Brush Border Membrane ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Cry Toxin

ABSTRACT Transgenic corn expressing the Bacillus thuringiensisCry1Ab gene is highly insecticidal to Ostrinia nubilalis(European corn borer) larvae. We ascertained whether Cry1F, Cry9C, or Cry9E recognizes the Cry1Ab binding site on the O. nubilalis brush border by three approaches. An optical biosensor technology based on surface plasmon resonance measured binding of brush border membrane vesicles (BBMV) injected over a surface of immobilized Cry toxin. Preincubation with Cry1Ab reduced BBMV binding to immobilized Cry1Ab, whereas preincubation with Cry1F, Cry9C, or Cry9E did not inhibit BBMV binding. BBMV binding to a Cry1F-coated surface was reduced when vesicles were preincubated in Cry1F or Cry1Ab but not Cry9C or Cry9E. A radioligand approach measured 125I-Cry1Ab toxin binding to BBMV in the presence of homologous (Cry1Ab) and heterologous (Cry1Ac, Cry1F, Cry9C, or Cry9E) toxins. Unlabeled Cry1Ac effectively competed for 125I-Cry1Ab binding in a manner comparable to Cry1Ab itself. Unlabeled Cry9C and Cry9E toxins did not inhibit 125I-Cry1Ab binding to BBMV. Cry1F inhibited125I-Cry1Ab binding at concentrations greater than 500 nM. Cry1F had low-level affinity for the Cry1Ab binding site. Ligand blot analysis identified Cry1Ab, Cry1Ac, and Cry1F binding proteins in BBMV. The major Cry1Ab signals on ligand blots were at 145 kDa and 154 kDa, but a strong signal was present at 220 kDa and a weak signal was present at 167 kDa. Cry1Ac and Cry1F binding proteins were detected at 220 and 154 kDa. Anti-Manduca sexta aminopeptidase serum recognized proteins of 145, 154, and 167 kDa, and anti-cadherin serum recognized the 220 kDa protein. We speculate that isoforms of aminopeptidase and cadherin in the brush border membrane serve as Cry1Ab, Cry1Ac, and Cry1F binding proteins.

scholarly journals Identification of cry1I-Type Genes from Bacillus thuringiensis Strains and Characterization of a Novel cry1I-Type Gene

2003 ◽  
Vol 69 (9) ◽  
pp. 5207-5211 ◽  
Author(s):  
Fuping Song ◽  
Jie Zhang ◽  
Aixing Gu ◽  
Yue Wu ◽  
Lanlan Han ◽  
...  
Keyword(s):  
Bacillus Thuringiensis ◽  
Diamondback Moth ◽  
Leaf Beetle ◽  
Beet Armyworm ◽  
Universal Primers ◽  
Cry Protein ◽  
Corn Borer ◽  
Pod Borer ◽  
Asian Corn Borer ◽  
Type Gene

ABSTRACT A PCR-restriction fragment length polymorphism method for identification of cry1I-type genes from Bacillus thuringiensis was established by designing a pair of universal primers based on the conserved regions of the genes to amplify 1,548-bp cry1I-type gene fragments. Amplification products were digested with the Bsp119I and BanI enzymes, and four kinds of known cry1I-type genes were successfully identified. The results showed that cry1I-type genes appeared in 95 of 115 B. thuringiensis isolates and 7 of 13 standard strains. A novel cry1I-type gene was found in one standard strain and six isolates. The novel cry1I gene was cloned from B. thuringiensis isolate Btc007 and subcloned into vector pET-21b. Then it was overexpressed in Escherichia coli BL21(DE3). The expressed product was shown to be toxic to the diamondback moth (Plutella xylostella), Asian corn borer (Ostrinia furnacalis), and soybean pod borer (Leguminivora glycinivorella). However, it was not toxic to the cotton bollworm (Helicoverpa armigera), beet armyworm (Spodoptera exigua), or elm leaf beetle (Pyrrhalta aenescens) in bioassays. Subsequently, the Cry protein encoded by this novel cry gene was designated Cry1Ie1 by the B. thuringiensis δ-endotoxin nomenclature committee.

scholarly journals Insecticidal Activity of Bacillus thuringiensis Proteins against Coleopteran Pests

Toxins ◽  
2020 ◽  
Vol 12 (7) ◽  
pp. 430 ◽  
Author(s):  
Mikel Domínguez-Arrizabalaga ◽  
Maite Villanueva ◽  
Baltasar Escriche ◽  
Carmen Ancín-Azpilicueta ◽  
Primitivo Caballero
Keyword(s):  
Bacillus Thuringiensis ◽  
Insecticidal Activity ◽  
Mode Of Action ◽  
Insect Species ◽  
Specific Information ◽  
Cry Proteins ◽  
Microbial Insecticide ◽  
Bt Toxins ◽  
Vip Proteins ◽  
Agro Forestry

Bacillus thuringiensis is the most successful microbial insecticide agent and its proteins have been studied for many years due to its toxicity against insects mainly belonging to the orders Lepidoptera, Diptera and Coleoptera, which are pests of agro-forestry and medical-veterinary interest. However, studies on the interactions between this bacterium and the insect species classified in the order Coleoptera are more limited when compared to other insect orders. To date, 45 Cry proteins, 2 Cyt proteins, 11 Vip proteins, and 2 Sip proteins have been reported with activity against coleopteran species. A number of these proteins have been successfully used in some insecticidal formulations and in the construction of transgenic crops to provide protection against main beetle pests. In this review, we provide an update on the activity of Bt toxins against coleopteran insects, as well as specific information about the structure and mode of action of coleopteran Bt proteins.

scholarly journals Structural, functional, and evolutionary analysis of Cry toxins of Bacillus thuringiensis: an in silico study

2021 ◽  
Vol 31 (1) ◽  
Author(s):  
Sujit Kumar Das ◽  
Sukanta Kumar Pradhan ◽  
Kailash Chandra Samal ◽  
Nihar Ranjan Singh
Keyword(s):  
Bacillus Thuringiensis ◽  
3D Structure ◽  
Evolutionary Relationship ◽  
Functional Domain ◽  
Main Body ◽  
Evolutionary Analysis ◽  
Structural Level ◽  
Cry Protein ◽  
Cry Proteins ◽  
Insect Order

Abstract Background Bacillus thuringiensis (Bt) is a gram-positive spore-forming soil bacterium that synthesizes crystalline (Cry) protein, which is toxic and causing pathogenicity against mainly three insect orders: Coleoptera, Diptera, and Lepidoptera. These crystalline protein inclusions, i.e., δ-endotoxins are successfully used as a bio-control agent against insect pests. Main body A total of 58 various Cry proteins belonging to these 3 insect orders were retrieved from SwissProt database and are categorized into different groups. Structural and functional analysis were performed to understand the functional domain arrangements at sequence level as well as at structural level involving both experimental and predicted 3-dimensional models. Besides, the analysis of evolutionary relationship involving all 58 observed Cry proteins at the sequence, domain, and structural levels were done using different bioinformatics tools. Evolutionary analysis revealed that some Cry proteins having toxicity for a specific insect order are found to be clustered for another different insect order, which concludes that they might have toxicity for more than one insect order. Three-dimensional (3D) structure analysis of both experimental and predicted models revealed that proteins might have toxicity for a specific insect order differ in their structural arrangements and was observed in Cry proteins belonging to 3 different insect orders. Conclusions It could be hypothesized that an inner-molecular domain shift or domain insertion/deletion might have taken place during the evolutionary process, which consequently causes structural and functional divergence of Bt. The study output may be helpful for understanding the diversity as well as specificity of the analyzed insecticidal proteins and their application as a biopesticide in the field of agriculture.

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

Toxins ◽  
2020 ◽  
Vol 12 (2) ◽  
pp. 133 ◽  
Author(s):  
Ayda Khorramnejad ◽  
Mikel Domínguez-Arrizabalaga ◽  
Primitivo Caballero ◽  
Baltasar Escriche ◽  
Yolanda Bel
Keyword(s):  
Bacillus Thuringiensis ◽  
Growth Phase ◽  
Brush Border ◽  
Brush Border Membrane ◽  
Insect Cells ◽  
3D Structure ◽  
Membrane Vesicles ◽  
Brush Border Membrane Vesicles ◽  
Host Susceptibility ◽  
Cry Proteins

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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