Transgenic crops producing insecticidal toxins from Bacillus thuringiensis (Bt) are commercially successful in reducing pest damage, yet knowledge of resistance mechanisms that threaten their sustainability is incomplete. Insect resistance to the pore-forming Cry1Ac toxin is correlated with the loss of high-affinity, irreversible binding to the mid-gut membrane, but the genetic factors responsible for this change have been elusive. Mutations in a 12-cadherin-domain protein confer some Cry1Ac resistance but do not block this toxin binding in in vitro assays. We sought to identify mutations in other genes that might be responsible for the loss of binding. We employed a map-based cloning approach using a series of backcrosses with 1,060 progeny to identify a resistance gene in the cotton pest Heliothis virescens that segregated independently from the cadherin mutation. We found an inactivating mutation of the ABC transporter ABCC2 that is genetically linked to Cry1Ac resistance and is correlated with loss of Cry1Ac binding to membrane vesicles. ABC proteins are integral membrane proteins with many functions, including export of toxic molecules from the cell, but have not been implicated in the mode of action of Bt toxins before. The reduction in toxin binding due to the inactivating mutation suggests that ABCC2 is involved in membrane integration of the toxin pore. Our findings suggest that ABC proteins may play a key role in the mode of action of Bt toxins and that ABC protein mutations can confer high levels of resistance that could threaten the continued utilization of Bt–expressing crops. However, such mutations may impose a physiological cost on resistant insects, by reducing export of other toxins such as plant secondary compounds from the cell. This weakness could be exploited to manage this mechanism of Bt resistance in the field.
Crystal toxin proteins from Bacillus thuringiensis (Bt) make ideal bioinsecticides because of their high potency against certain insects and lack of activity against most other species. Transgenic cotton and maize expressing pore-forming Cry1A Bt-toxins are now widely used in agriculture, enabling substantial reductions in the use of chemical insecticides. However this has greatly increased the selection pressure in pest populations for toxin resistance. Preventing or delaying the development of this resistance is a high priority, to avoid a replay of the onset of insecticide resistance brought on by dependency on chemical pesticides. Because the molecular details of Bt mode of action are still not fully understood, insect strains collected from the field and selected to high levels of resistance in the laboratory are useful in discovering the obstacles the toxin must overcome before it finally forms the pore and kills the insect. We used a genetic approach to explore a poorly understood step in the toxin mode of action, which is blocked in an extremely resistant strain of an important cotton pest. As well as providing the tools to diagnose this type of resistance when it appears in the field, this discovery suggests factors that may counteract its eventual spread.