Mutations in Promoter Region Enhanced PxαE27 -Mediated β-Cypermethrin Resistance in Plutella xylostella

Insecticide resistance in insects often involves overexpression of carboxylesterase (CarE) genes. In the diamondback moth, Plutella xylostella, β-cypermethrin resistance is linked to elevated expressions of PxαE9, PxαE27, and PxαE14 . To elucidate the underlying transcriptional regulation, we identified their core promoters and predicted, via in silico analysis (ALGGEN and JASPAR), that the transcription factor Dfd binds to PxαE27 and PxαE14 . Experimental validation using dual-luciferase assays, site-directed mutagenesis, yeast one-hybrid assays, and RNA interference confirmed that Dfd enhances PxαE27 transcription by direct promoter binding. Furthermore, a two-nucleotide mutation in the PxαE27 promoter of a field-resistant population created a novel Dfd-binding site, further upregulating expression and increasing β-cypermethrin resistance. This study reveals how cis -regulatory mutations recruit specific transcription factors to elevate CarE expression and drive insecticide resistance, providing a mechanistic basis for potential resistance management strategies.