Members of the insect ATP binding cassette transporter subfamily C2 (ABCC2) in several moth species are known as receptors for the Cry1Ac insecticidal protein from Bacillus thuringiensis (Bt). Mutations that abolish the functional domains of ABCC2 are known to cause resistance to Cry1Ac, although the reported levels of resistance vary widely depending on insect species. In this study, the function of the ABCC2 gene as a putative Cry1Ac receptor in Helicoverpa zea, a major pest of over 300 crops, was evaluated using CRISPR/Cas9 to progressively eliminate different functional ABCC2 domains. Results from bioassays with edited insect lines support that mutations in ABCC2 were associated with Cry1Ac resistance ratios (RR) ranging from 7.3- to 39.8-fold. No significant differences in susceptibility to Cry1Ac were detected between H. zea with partial or complete ABCC2 knockout, although the highest levels of tolerance were observed when knocking out half of ABCC2. Based on >500-1000-fold RRs reported in similar studies for closely related moth species, the low RRs observed in H. zea knockouts support that ABCC2 is not a major Cry1Ac receptor in this insect.
Keywords: ABCC2; ATP binding cassette transporter; Bt toxin; CRISPR; Cry1A; Helicoverpa zea; bollworm; functional genomics; genome editing; knockout; resistance.