Lmx1b is essential for molting and leg integrity in Henosepilachna vigintioctopunctata: Implications for RNA biopesticide development.

LIM homeodomain transcription factor 1 beta (Lmx1b), a member of the LIM-homeodomain family, plays essential roles in tissue differentiation and organ development in diverse eukaryotes, yet its biological functions in insects remain largely unknown. Here, we identify Lmx1b as a highly effective RNA interference (RNAi) target in the 28-spotted ladybeetle, Henosepilachna vigintioctopunctata, a serious solanaceous pest. RNAi-mediated silencing of HvLmx1b prevented larval molting and pupation, resulting in complete mortality. HvLmx1b knockdown markedly reduced endogenous 20-hydroxyecdysone (20E) titers, suppressed Halloween genes and the ecdysone signaling pathway, and downregulated multiple genes involved in chitin metabolism and cuticle tanning, resulting in defective cuticle formation. Ultrastructural analyses further revealed progressive degeneration of epidermal cells and leg muscles, indicating an essential role for HvLmx1b in maintaining tissue integrity during postembryonic development. In silico off-target analysis against representative non-target arthropods and beneficial organisms predicted a low risk of unintended gene silencing by dsHvLmx1b. Collectively, our findings establish HvLmx1b as an upstream developmental regulator that coordinates ecdysteroid signaling, cuticle formation and postembryonic tissue homeostasis, and highlight its potential as an RNAi target for management of H. vigintioctopunctata.