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Lepidopteran midgut aminopeptidases N (APNs) are phylogenetically divided into eight clusters, designated as APN1–8. Although APN1 has been implicated as one of the receptors for Cry1Ac in several species, its potential role in the mode of action of Cry2Ab has not been functionally determined so far. To test whether APN1 also acts as one of the receptors for Cry1Ac in  Helicoverpa zea  and even for Cry2Ab in this species, we conducted a gain of function analysis by heterologously expressing  H. zea  APN1 ( Hz APN1) in the midgut and fat body cell lines of  H. zea  and the ovarian cell line of  Spodoptera frugiperda  (Sf9) and a loss of function analysis by RNAi (RNA interference) silencing of the endogenous APN1 in the three cell lines using the  HzAPN1  double strand RNA (dsRNA). Heterologous expression of  HzAPN1  significantly increased the susceptibility of the three cell lines to Cry1Ac, but had no effects on their susceptibility to Cry2Ab. Knocking down of the endogenous APN1 made the three cell lines resistant to Cry1Ac, but didn’t change cell lines susceptibility to Cry2Ab. The findings from this study demonstrate that HzAPN1 is a functional receptor of Cry1Ac, but not Cry2Ab.

APN1 is a functional receptor of Cry1Ac but not Cry2Ab in Helicoverpa zea