Cell-cycle-dependent resistance to<i>Bacillus thuringiensis</i>Cry1C toxin in Sf9 cells

Avisar, Dror; Segal, Michal; Sneh, B.; Zilberstein, Aviah

doi:10.1242/jcs.02440

Cited by 15 publications

(13 citation statements)

References 63 publications

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“…Several studies suggested that lipid rafts might play a central role in Bt-toxins toxicity. For example, the presence of lipid rafts was shown to be essential for the toxicity of Cry1C on Sf9 Lepidoptera cells [27]. However, the specific proteins associated with this mechanism have not been identified yet.…”

Section: Discussionmentioning

confidence: 99%

Gene expression patterns and sequence polymorphisms associated with mosquito resistance to Bacillus thuringiensis israelensis toxins

et al. 2014

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BackgroundDespite the intensive use of Bacillus thuringiensis israelensis (Bti) toxins for mosquito control, little is known about the long term effect of exposure to this cocktail of toxins on target mosquito populations. In contrast to the many cases of resistance to Bacillus thuringiensis Cry toxins observed in other insects, there is no evidence so far for Bti resistance evolution in field mosquito populations. High fitness costs measured in a Bti selected mosquito laboratory strain suggest that evolving resistance to Bti is costly. The aim of the present study was to identify transcription level and polymorphism variations associated with resistance to Bti toxins in the dengue vector Aedes aegypti. We used RNA sequencing (RNA-seq) for comparing a laboratory-selected strain showing elevated resistance to Bti toxins and its parental non-selected susceptible strain. As the resistant strain displayed two marked larval development phenotypes (slow and normal), each phenotype was analyzed separately in order to evidence potential links between resistance mechanisms and mosquito life-history traits.ResultsA total of 12,458 genes were detected of which 844 were differentially transcribed between the resistant and susceptible strains. Polymorphism analysis revealed a total of 68,541 SNPs of which 12,571 SNPs exhibited more than 40% frequency difference between the resistant and susceptible strains, affecting 2,953 genes. Bti resistance is associated with changes in the transcription level of enzymes involved in detoxification and chitin metabolism. Among previously described Bti-toxin receptors, four alkaline phosphatases (ALPs) were differentially transcribed between resistant and susceptible larvae, and non-synonymous changes affected the protein sequence of one cadherin, six aminopeptidases (APNs) and four α-amylases. Other putative Cry receptors located in lipid rafts, such as flotillin and glycoside hydrolases, were under-transcribed and/or contained non-synonymous substitutions. Finally, immunity-related genes showed contrasted transcription and polymorphisms patterns between the two developmental resistant phenotypes, suggesting the existence of trade-offs between Bti-resistance, life-history traits and immunity.ConclusionsThe present study is the first to analyze the whole transcriptome of Bti-resistant mosquitoes by RNA-seq, shedding light on the importance of studying both transcription levels and sequence polymorphism variations to get a comprehensive view of insecticide resistance.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2164-15-926) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Gene expression patterns and sequence polymorphisms associated with mosquito resistance to Bacillus thuringiensis israelensis toxins

et al. 2014

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“…The presence of lipid rafts in the DRM fraction provided an approach to bridge the model that Cry1 toxins insert into lipid rafts 9–11 to mosquitocidal Cry toxins. The detection of membrane-bound Cry4Ba toxin in the DRM fraction (Fig.…”

Section: Discussionmentioning

confidence: 99%

Proteome Analysis of Cry4Ba Toxin-interacting Aedes aegypti Lipid Rafts using geLC–MS/MS

et al. 2012

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Lipid rafts are microdomains in the plasma membrane of eukaryotic cells. Among their many functions, lipid rafts are involved in cell toxicity caused by pore forming bacterial toxins including Bacillus thuringiensis (Bt) Cry toxins. We isolated lipid rafts from brush border membrane vesicles (BBMV) of Aedes aegypti larvae as a detergent resistant membrane (DRM) fraction on density gradients. Cholesterol, aminopeptidase (APN), alkaline phosphatase (ALP) and the raft marker flotillin were preferentially partitioned into the lipid raft fraction. When mosquitocidal Cry4Ba toxin was pre-incubated with BBMV, Cry4Ba localized to lipid rafts. A proteomic approach based on one dimensional gel electrophoresis, in-gel trypsin digestion, followed by liquid chromatography-mass spectrometry (geLC-MS/MS) identified a total of 386 proteins. Of which many are typical lipid raft marker proteins including flotillins and glycosylphosphatidylinositol (GPI)-anchored proteins. Identified raft proteins were annotated in silico for functional and physicochemical characteristics. Parameters such as distribution of isoelectric point, molecular mass, and predicted post-translational modifications relevant to lipid raft proteins (GPI anchorage and myristoylation or palmitoylation) were analyzed for identified proteins in the DRM fraction. From a functional point of view, this study identified proteins implicated in Cry toxin interactions as well as membrane-associated proteins expressed in the mosquito midgut that have potential relevance to mosquito biology and vector management.

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“…To investigate further the role of lipid rafts in JcDNV endocytosis, we took advantage of the conservation of the caveolin-1 protein in Lepidoptera and we used the fluorescent cholera toxin subunit B (FITC-CTxB) as a marker of these microdomains (24,30). Our results indicate that, 10 min after its addition to the luminal side of the epithelium, FITC-CTxB colocalized, although partially, with caveolin-1-positive dots, validating both markers for studying this pathway (see Fig.…”

Section: Jcdnv Particles Reach the Midgut Cells Within 15 Min After Imentioning

confidence: 99%

Densovirus Crosses the Insect Midgut by Transcytosis and Disturbs the Epithelial Barrier Function

Wang

Gosselin-Grenet

Castelli

et al. 2013

J Virol

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g Densoviruses are parvoviruses that can be lethal for insects of different orders at larval stages. Although the horizontal transmission mechanisms are poorly known, densoviral pathogenesis usually starts with the ingestion of contaminated food by the host. Depending on the virus, this leads to replication restricted to the midgut or excluding it. In both cases the success of infection depends on the virus capacity to enter the intestinal epithelium. Using the Junonia coenia densovirus (JcDNV) as the prototype virus and the lepidopteran host Spodoptera frugiperda as an interaction model, we focused on the early mechanisms of infection during which JcDNV crosses the intestinal epithelium to reach and replicate in underlying target tissues. We studied the kinetics of interaction of JcDNV with the midgut epithelium and the transport mechanisms involved. Using several approaches, in vivo, ex vivo, and in vitro, at molecular and cellular levels, we show that JcDNV is specifically internalized by endocytosis in absorptive cells and then crosses the epithelium by transcytosis. As a consequence, viral entry disturbs the midgut function. Finally, we showed that four mutations on the capsid of JcDNV affect specific recognition by the epithelial cells but not their binding.

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Cell-cycle-dependent resistance toBacillus thuringiensisCry1C toxin in Sf9 cells

Cited by 15 publications

References 63 publications

Gene expression patterns and sequence polymorphisms associated with mosquito resistance to Bacillus thuringiensis israelensis toxins

Gene expression patterns and sequence polymorphisms associated with mosquito resistance to Bacillus thuringiensis israelensis toxins

Proteome Analysis of Cry4Ba Toxin-interacting Aedes aegypti Lipid Rafts using geLC–MS/MS

Densovirus Crosses the Insect Midgut by Transcytosis and Disturbs the Epithelial Barrier Function

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