Abstract
Background Bacillus thuringiensis Cry toxins, that are used worldwide in insect control, kill insects by a mechanism that depends on their ability to form oligomeric pores that insert into the insect-midgut cells. These toxins are being used worldwide in transgenic plants or spray to control insect pests in agriculture. However, a major concern has been the possible effects of these insecticidal proteins on non-target organisms mainly in ecosystems adjacent to agricultural fields.Methodology/Principal FindingsWe isolated and characterized 11 non-toxic mutants of Cry1Ab toxin affected in different steps of the mechanism of action namely binding to receptors, oligomerization and pore-formation. These mutant toxins were analyzed for their capacity to block wild type toxin activity, presenting a dominant negative phenotype. The dominant negative phenotype was analyzed at two levels, in vivo by toxicity bioassays against susceptible Manduca sexta larvae and in vitro by pore formation activity in black lipid bilayers. We demonstrate that some mutations located in helix α-4 completely block the wild type toxin activity at sub-stoichiometric level confirming a dominant negative phenotype, thereby functioning as potent antitoxins.Conclusions/SignificanceThis is the first reported case of a Cry toxin dominant inhibitor. These data demonstrate that oligomerization is a fundamental step in Cry toxin action and represent a potential mechanism to protect special ecosystems from the possible effect of Cry toxins on non-target organisms.
Highlights
Bacillus thuringiensis (Bt) bacteria produce crystal proteins that have insecticidal activity
Dominant negative (DN) inhibitors which are inactive mutant-toxins, able to form oligomer structures but affected in their pore formation activity, work as powerful inhibitors since they are able to co-assemble into hetero-oligomers together with the wild type toxin resulting in an effective inactivation of pore formation and toxicity [14,15,16]
Cry1Ab mutant characterization We isolated and characterized Cry1Ab mutants affected at different steps of their mode of action, namely receptor binding, oligomerization and pore-formation to determine if any of them showed a dominant negative (DN) phenotype
Summary
Bacillus thuringiensis (Bt) bacteria produce crystal proteins (denominated Cry toxins) that have insecticidal activity. There are still concerns related to the possible impact of by products from transgenic Bt crops as Bt-cotton and Bt-corn on non-target organisms in ecosystems adjacent to agricultural fields [5,6,7,8,9]. Bacillus thuringiensis Cry toxins, that are used worldwide in insect control, kill insects by a mechanism that depends on their ability to form oligomeric pores that insert into the insect-midgut cells. These toxins are being used worldwide in transgenic plants or spray to control insect pests in agriculture. A major concern has been the possible effects of these insecticidal proteins on non-target organisms mainly in ecosystems adjacent to agricultural fields
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have