Fig 3. Osy function prevents penetration of xenobiotics.

In dye penetration assays, the 1^st instar wild-type (A) and osy mutant (osy^ΔMiM18, B) larvae do not take up Eosin Y at 25°C Down-regulation of osy transcripts by epidermal RNAi (osy^epIR, 69B-Gal4 x UAS-KK109988, C) or ubiquitous (osy^ubIR, da/7063-Gal4 x UAS-KK109988, D) RNAi does not cause dye penetration at 25°C in 1^st instar larvae. At 40°C, the cuticle of wild-type 1^st instar larvae is impermeable to Eosin Y (D), while in osy mutant 1^st instar larvae the dye penetrates the cuticle (E). At 25°C, in 1^st instar larvae with reduced osy transcript levels in the whole body (F) or in the epidermis (G), Eosin Y does not penetrate the cuticle. The cuticle of 1^st instar larvae with reduced osy transcript levels in the whole body (H) or in the epidermis (I) fails to withstand dye penetration at 40°C. The envelope (arrow) of the wild-type 1^st instar larva auto-fluoresces upon excitation with UV light (I). Envelope auto-fluorescence is unchanged in osy^ΔMiM18 1^st instar larvae (J). By contrast, envelope auto-fluorescence is reduced in snu mutant 1^st instar larvae (K). This defect allows Eosin Y penetration into these larvae at 25°C (inset). The envelope (env) of wild-type stage 17 ready-to-hatch embryos consists of alternating electron-lucid and electron-dense sheets in electron-micrographs (L). The envelope of stage 17 ready-to-hatch osy^ΔMiM18 embryos is normal in electron-micrographs (M). The asterisks (*) mark unspecific material at the surface of the cuticle. vit vitelline membrane. Scale bar 500nm.