Coordinated P450–UGT detoxification contributes to furanocoumarin tolerance associated with host plant range divergence in Helicoverpa

Plants and herbivorous insects are engaged in a continuous evolutionary arms race driven by chemical defences and counter-defences. How insects integrate distinct detoxification systems to overcome diverse phytochemicals, and how such mechanisms contribute to host-range divergence, remains poorly understood. Here, we uncover a cooperative cytochrome P450-UDP-glycosyltransferase (UGT) cascade that mediates furanocoumarin tolerance in Helicoverpa armigera. Comparative genomic analyses across Helicoverpa and related noctuid species revealed that the UGT33 family is the most extensively expanded and dynamically diversified UGT lineage in Helicoverpa. Targeted CRISPR-Cas9 knockouts of UGT33 gene clusters in polyphagous H. armigera demonstrated their essential roles in detoxifying the furanocoumarins xanthotoxin and imperatorin. Combined metabolic and functional assays further established a sequential detoxification pathway, in which the cytochrome P450 CYP6AE19 catalyses either the O-dealkylation of xanthotoxin to yield xanthotoxol or the aromatic-carbon hydroxylation of xanthotoxin to form 5-hydroxyxanthotoxin, which are subsequently glycosylated by UGT33 enzymes to form their less-toxic glucosides. In contrast, impaired CYP6AE19-UGT33 coordination in the oligophagous Helicoverpa assulta was associated with reduced xanthotoxin detoxification and high sensitivity to this compound. Together, these findings provide direct evidence for coordinated Phase I-Phase II detoxification of a plant defensive compound in insects and show that functional divergence in this pathway contributes to interspecific differences in plant toxin tolerance, which are associated with contrasting dietary breadth in closely related herbivorous insects.