Insect Resistance to Bacillus thuringiensis Toxin Cry2Ab Is Conferred by Mutations in an ABC Transporter Subfamily A Protein

Abstract

The use of conventional chemical insecticides and bacterial toxins to control lepidopteran pests of global agriculture has imposed significant selection pressure leading to the rapid evolution of insecticide resistance. Transgenic crops (e.g., cotton) expressing the Bt Cry toxins are now used world wide to control these pests, including the highly polyphagous and invasive cotton bollworm Helicoverpa armigera. Since 2004, the Cry2Ab toxin has become widely used for controlling H. armigera, often used in combination with Cry1Ac to delay resistance evolution. Isolation of H. armigera and H. punctigera individuals heterozygous for Cry2Ab resistance in 2002 and 2004, respectively, allowed aspects of Cry2Ab resistance (level, fitness costs, genetic dominance, complementation tests) to be characterised in both species. However, the gene identity and genetic changes conferring this resistance were unknown, as was the detailed Cry2Ab mode of action. No cross-resistance to Cry1Ac was observed in mutant lines. Biphasic linkage analysis of a Cry2Ab-resistant H. armigera family followed by exon-primed intron-crossing (EPIC) marker mapping and candidate gene sequencing identified three independent resistance-associated INDEL mutations in an ATP-Binding Cassette (ABC) transporter gene we named HaABCA2. A deletion mutation was also identified in the H. punctigera homolog from the resistant line. All mutations truncate the ABCA2 protein. Isolation of further Cry2Ab resistance alleles in the same gene from field H. armigera populations indicates unequal resistance allele frequencies and the potential for Bt resistance evolution. Identification of the gene involved in resistance as an ABC transporter of the A subfamily adds to the body of evidence on the crucial role this gene family plays in the mode of action of the Bt Cry toxins. The structural differences between the ABCA2, and that of the C subfamily required for Cry1Ac toxicity, indicate differences in the detailed mode-of-action of the two Bt Cry toxins.

A P-Glycoprotein Is Linked to Resistance to the Bacillus thuringiensis Cry3Aa Toxin in a Leaf Beetle

“…Expression of other Bt-toxin binding proteins such as aminopeptidase [47,48,49] or cadherin [18,19] has a much weaker effect. Recently, several mutations in ABCA2, a member of the A subfamily of ABC proteins, were found to confer high resistance against Cry2Ab1 in two Lepidoptera, Helicoverpa armigera and H. punctigera [13]. Our study adds a third subfamily of ABC proteins and a different toxin, active against Coleoptera but not Lepidoptera, to this interaction.…”

“…A previous report indicated that, in C. tremula , there was no difference in sequence and in expression of three APNs between insects of the susceptible and the resistant strains [10]. We turned to ABC proteins because of their association with resistance to Cry1A and Cry2A toxins in Lepidoptera [11,12,13,14]. We used a recent analysis of the tissue-specific expression of genes encoding ABC proteins in C. populi , a sister species of C. tremula , to identify ABC proteins expressed in the larval midgut.…”

Specificity determinants for Cry insecticidal proteins: Insights from their mode of action

“…There has been much interest recently in the interaction of Cry toxins with members of the ATP binding cassette (ABC) protein family, particularly subfamilies C2 and A. Alterations in these ABC proteins are linked with resistance against Cry1 toxins (Heckel, 2012;Park et al, 2014;Xiao et al, 2014) and Cry2Ab (Tay et al, 2015). Although the exact role of these proteins remains unclear it has been proposed that they may act as receptors but only when in a particular conformational state (Heckel, 2012) …”

Functional characterization of Aedes aegypti alkaline phosphatase ALP1 involved in the toxicity of Cry toxins from Bacillus thuringiensis subsp. israelensis and jegathesan

“…In lepidopteran insects, four major Cry1A toxin protein receptors have been identified – cadherins, ABCC transporters, aminopeptidases (APNs) and alkaline phosphatases (ALPs) (Atsumi et al 2012; Bravo et al 2011; Gahan et al 2010; Pardo-Lopez et al 2012; Pigott and Ellar 2007; Soberon et al 2009; Tay et al 2015). APN and ALP are thought to be receptors that facilitate localization of toxin monomers to midgut microvilli before toxin interaction with cadherin.…”