The wing-deficient mutant, flügellos (fl), of the silkworm, lacks four wings in the pupa and the adult. Previous studies have suggested that the fl wing discs lose responsiveness to ecdysteroid during metamorphosis. To test this hypothesis at the molecular level we compared the expression of 12 genes when the wing discs from the wild-type (WT) and fl larvae were cultured in the presence or absence of 20-hydroxyecdysone (20E). Most of the genes tested here, ecdysteroid-inducible (EcR-A, -B1, and E75) and noninducible genes (actin A3, beta-tubulin, apterous (ap), USP, and BHR38) were normally expressed in the fl wing discs. However, the amounts of mRNAs of two ecdysteroid-inducible genes, BHR3 (early-late gene; Bombyx homologue to DHR3 and MHR3) and Urbain (wing-specific late gene), were reduced to about 50% and 20% of WT in the cultured fl wing discs, respectively. We analyzed developmental profiles of these mRNAs during metamorphosis. They also demonstrated decreased BHR3 and Urbain mRNA 2 days after the onset of wandering. This reduction in transcription of BHR3 in the fl mutant was observed only in the wing disc, not in the testis and fatbody. These results imply that the aberrant expression of the fl gene affects the downstream pathway of ecdysteroid signaling specifically in the wing discs and thus leads to a deficiency in wing formation.