Prevention of desiccation is a constant challenge for terrestrial organisms. Land insects have an extracellular coat, the cuticle, that plays a major role in protection against exaggerated water loss. Here, we report that the ABC transporter Oskyddad (Osy)—a human ABCA12 paralog—contributes to the waterproof barrier function of the cuticle in the fruit fly Drosophila melanogaster. We show that the reduction or elimination of Osy function provokes rapid desiccation. Osy is also involved in defining the inward barrier against xenobiotics penetration. Consistently, the amounts of cuticular hydrocarbons that are involved in cuticle impermeability decrease markedly when Osy activity is reduced. GFP-tagged Osy localises to membrane nano-protrusions within the cuticle, likely pore canals. This suggests that Osy is mediating the transport of cuticular hydrocarbons (CHC) through the pore canals to the cuticle surface. The envelope, which is the outermost cuticle layer constituting the main barrier, is unaffected in osy mutant larvae. This contrasts with the function of Snu, another ABC transporter needed for the construction of the cuticular inward and outward barriers, that nevertheless is implicated in CHC deposition. Hence, Osy and Snu have overlapping and independent roles to establish cuticular resistance against transpiration and xenobiotic penetration. The osy deficient phenotype parallels the phenotype of Harlequin ichthyosis caused by mutations in the human abca12 gene. Thus, it seems that the cellular and molecular mechanisms of lipid barrier assembly in the skin are conserved during evolution. As in humans, lipids on the surface of the skin of insects protect the organism against excessive water loss and penetration of potentially harmful substances. During evolution, a greasy surface was indeed an essential trait for adaptation to life outside a watery environment. Here, we show that the membrane-gate transporter Oskyddad (Osy) is needed for the deposition of barrier lipids on the integument surface in the fruit fly Drosophila melanogaster through extracellular nano-tubes, called pore canals. In principle, the involvement of Osy parallels the scenario in humans, where the membrane-gate transporter ABCA12 is implicated in the construction of the lipid-based stratum corneum of the skin. In both cases, mutations in the genes coding for the respective transporter cause rapid water-loss and are lethal soon after birth. We conclude that the interaction between the organism and the environment obviously implies an analogous mechanism of barrier formation and function in vertebrates and invertebrates.

The resistance of pests to common insecticides is a global issue that threatens food production worldwide. Diamide insecticides target insect ryanodine receptors (RyRs), causing uncontrolled calcium release from the sarcoplasmic and endoplasmic reticulum. Despite their high potency and species selectivity, several resistance mutations have emerged. Using a chimeric RyR (chiRyR) approach and cryo-electron microscopy (cryo-EM), we investigate how insect RyRs engage two different diamide insecticides from separate families: flubendiamide, a phthalic acid derivative, and tetraniliprole, an anthranilic compound. Both compounds target the same site in the transmembrane region of the RyR, albeit with different poses, and promote channel opening through coupling with the pore-forming domain. To explore the resistance mechanisms, we also solve two cryo-EM structures of chiRyR carrying the two most common resistance mutations, I4790M and G4946E, both alone and in complex with the diamide insecticide chlorantraniliprole. The resistance mutations perturb the local structure, directly reducing the binding affinity and altering the binding pose. Our findings elucidate the mode of action of different diamide insecticides, reveal the molecular mechanism of resistance mutations, and provide important clues for the development of novel pesticides that can bypass the resistance mutations.