Fig. 3. A transient high amount of Ubx protein permanently represses the endogenous Ubx gene. (A-A''') Haltere disc of an ap-Gal4 UAS-GFP/+; UAS-Ubx Ubx^lac1/+ larva, grown at 17°C. GFP (A, in green) and high Ubx protein expression (A', in red) are observed in the dorsal compartment, whereas the expression of the Ubx^lac1 reporter (A'', greyscale) is absent in that region (arrow). A''', merged image. In these and subsequent panels, d and v stand for dorsal and ventral compartments, respectively. (B-D') ap-Gal4 UASGFP/+; UAS-Ubx Ubx^lac1/tub-Gal80^ts haltere discs from larvae grown for 24-48 hours at 17°C, kept for 3 (B,D) or 2 (C) days at 29°C, and then transferred to 17°C for 3 (B, 3d) or 6 (C,D, 6d) days. Note that GFP (in green) and total Ubx (in red) protein expression progressively disappear and that the Endo-Ubx expression (D,D', greyscale and blue) is not restored. (E,F) Comparison of Ubx expression in ap-Gal4 UAS-GFP/+; Df109 UAS-dsUbx/tub-Gal80^ts haltere discs from larvae grown for 24-48 hours at 17°C and then at 29°C (E), or reared for 24-48 hours at 17°C, 3 days at 29°C and 4 days (4d) at 17°C (F). The absence of Ubx protein by repression of transcription (B-D) is permanent, whereas the absence by RNA interference allows the recovery of Ubx expression (F). (G) In ap-Gal4/UAS-Ubx flies, the halteres are greatly reduced (arrow). (H) By contrast, in ap-Gal4/UAS-Ubx; tub-Gal80^ts/+ adults that were transferred to 17°C after being at 29°C for 2 days during the second and early third larval instars, halteres are strongly transformed into wings (arrow). (I-L) Clones marked by the absence of GFP (in green, I), which expressed Ubx-HA transiently, induced in larvae of the hs-flp; UAS-Ubx-HA/tub-Gal80^ts; Ubx^lac1/tub>Ubi-GFP, y⁺>Gal4 genotype. Within the clone, exogenous Ubx protein is no longer present (J, in red) and the endogenous Ubx expression is continuously repressed (K, greyscale). Merged image in L. Discs are oriented with the anterior compartment towards the left and the ventral compartment upwards.