Proteomic analysis of Cry2Aa-binding proteins and their receptor function in Spodoptera exigua

Lin Qiu,Chaoliang Lei,Boyao Zhang,Weihua Ma,Lang Liu,Lizhen Chen,Xiaoping Wang

doi:10.1038/srep40222

Abstract

The bacterium Bacillus thuringiensis produces Crystal (Cry) proteins that are toxic to a diverse range of insects. Transgenic crops that produce Bt Cry proteins are grown worldwide because of their improved resistance to insect pests. Although Bt “pyramid” cotton that produces both Cry1A and Cry2A is predicted to be more resistant to several lepidopteran pests, including Spodoptera exigua, than plants that produce Cry1Ac alone, the mechanisms responsible for the toxicity of Cry2Aa in S. exigua are not well understood. We identified several proteins that bind Cry2Aa (polycalin, V-ATPase subunits A and B, actin, 4-hydroxybutyrate CoA-transferase [4-HB-CoAT]), and a receptor for activated protein kinase C (Rack), in S. exigua. Recombinant, expressed versions of these proteins were able to bind the Cry2Aa toxin in vitro assays. RNA interference gene knockdown of the Se-V-ATPase subunit B significantly decreased the susceptibility of S. exigua larvae to Cry2Aa, whereas knockdown of the other putative binding proteins did not. Moreover, an in vitro homologous competition assay demonstrated that the Se-V-ATPase subunit B binds specifically to the Cry2Aa toxin, suggesting that this protein acts as a functional receptor of Cry2Aa in S. exigua. This the first Cry2Aa toxin receptor identified in S. exigua brush-border membrane vesicles.

Highlights

The bacterium Bacillus thuringiensis produces Crystal (Cry) proteins that are toxic to a diverse range of insects
The Crystal (Cry) toxins produced by Bacillus thuringiensis (Bt) are a diverse group of proteins that are used to control a broad range of insect pests[1]
We present the first analysis of Cry2Aa receptor proteins in S. exigua brush-border membrane vesicles (BBMVs)

Summary

Introduction

The bacterium Bacillus thuringiensis produces Crystal (Cry) proteins that are toxic to a diverse range of insects. An in vitro homologous competition assay demonstrated that the Se-V-ATPase subunit B binds to the Cry2Aa toxin, suggesting that this protein acts as a functional receptor of Cry2Aa in S. exigua. Transgenic plants that express both Cry1Ac and Cry2Ab toxin would be expected to be much more resistant to lepidopteran pests, especially the beet armyworm Spodoptera exigua[3,14]. Some studies suggest that S. exigua is less sensitive to Cry2Aa/b than to Cry1B, Cry1C or other toxins[18,19], Bt crops producing both Cry1Ac and Cry2Aa/b (Cry2Ab in the case of cotton) are predicted to be more resistant to S. exigua, and several other lepidopteran pests, than those currently cultivated in China which produce only Cry1Ac3,15,20–22.

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