Functional characterization of Niemann–Pick proteins type C2 as odorant carriers in <i>Neoseiulus barkeri</i> olfactory perception

Arthropods utilize olfactory function to detect informational substances within their natural habitats, thereby facilitating essential life activities such as foraging, mating, and oviposition. Niemann-Pick protein type C2 (NPC2), a putative odorant carrier, has not been fully described in the olfactory perception of predatory mites. In this study, we characterized in detail the olfactory perception functions of NbNPC2-2 and NbNPC2-3 in the important predatory mite Neoseiulus barkeri. The results show that 2 NPC2 proteins, with 6 conserved cysteine sites, are significantly upregulated (by 2.93-fold for NbNPC2-2 and 1.66-fold for NbNPC2-3) with methyl salicylate (MeSA), an important herbivore-induced plant volatile (HIPV). Knocking down both NbNPC2-2 and NbNPC2-3 prevented N. barkeri from preferring pest-infested plants, and reduced their crawling speed to 2.92 and 3.12 mm/s under MeSA treatment. The affinity analysis revealed that NPC2-2 binds to HIPVs β-ionone, MeSA, and ocimene (Ki = 28.55 ± 2.12, 40.73 ± 3.89, and 45.15 ± 2.22, respectively), and NPC2-3 binds to 4-ethylbenzaldehyde (Ki = 51.67 ± 0.23). Molecular docking further confirmed hydrogen bonding interactions between NPC2-2, NPC2-3, and these HIPVs. These results indicated that NPC2-2 and NPC2-3 act as odor carriers involved in the olfactory perception of mites toward HIPVs. This enhances understanding of the molecular mechanism of phytoseiid olfaction.