RNAi-mediated knockdown of <i>fruitless</i> in <i>Plutella xylostella</i> (Lepidoptera: Plutellidae) disrupts female sex pheromone biosynthesis and male courtship behavior

Strong fecundity is an important reason why the diamondback moth, Plutella xylostella (L.) (Lepidoptera: Plutellidae), has become one of the most serious pests of cruciferous vegetables worldwide. Disrupting with courtship and mating behaviors has emerged as an important strategy for insect management. The fruitless (fru) gene encodes a transcription factor that contains a BTB (Broad-Complex, Tramtrack and Bric a brac) and a zinc finger pair related to the C2H2 class. It plays a crucial role in regulating insect courtship behavior. In this study, the fru gene of P. xylostella (Pxfru) was cloned, revealing 7 alternative splicing forms (Pxfru-1 to Pxfru-7). Pxfru-1 to Pxfru-3 were non-sex-specific transcripts, while the remaining forms were male-specific. Subcellular localization experiments demonstrated that the transcripts encoding proteins containing BTB and zinc finger domains (Pxfru-1 to Pxfru-3) localized to the cell nucleus, whereas Pxfru-4 and Pxfru-5, which contain only one BTB domain, were localized in the nucleus and cytoplasm, respectively. Knockdown the expression of fru in male moths delayed occurrence of mating and reduced their preference to female sex pheromones. Meanwhile, suppression of fru expression in female P. xylostella decreased their attractiveness to males. The results of GC-MS and Y-tube olfaction experiments indicated that this change may be attributed to alterations in the proportion of sex pheromones. This study represents the first report of the fru gene influencing pheromone ratios in female insects, and provides a new perspective for understanding the function of fru in the courtship behavior of non-model insects.