Fig. 5. rdx regulates Ci behind the MF. Third instar eye-antennal imaginal discs (B-F) are oriented with dorsal upwards, anterior towards the right, an arrowhead marking the MF and an asterisk marking the ocellar region of the head capsule. A schematic (A) shows Ci155 levels (red) across the disc, with the Hh-secreting photoreceptors posterior to the MF in grey. Anterior to the MF, in the absence of Hh, Ci155 is processed via Cul1 (blue arrow) to CiR. Posterior to the MF, Ci155 is degraded (red blobs) via Cul3 (green arrow). (B) rdx as detected by ß-gal activity (blue) in rdx²/+ discs (rdxZ), is found posterior to the MF, in the ocellar region, and at the compartment border of the antennal portion (arrow). At higher resolution (C), nuclear-localized ß-gal protein in rdxZ (green) co-localized with Elav (not shown) in all photoreceptors and cone cells, while Ci155 (red) accumulated in the intervening cells. ß-Gal protein is also found in many basal nuclei, corresponding to the position of lattice cell nuclei (not shown). (D,E) rdx clones are marked by loss of GFP (green), while Ci155 is in red. The three arrows (left to right) indicate, respectively, clones behind, within and infront of the MF. The hypomorphic allele rdx¹ was used as a control to illustrate normal patterning (D). In clones of the loss-of-function allele rdx⁵ (E), Ci155 accumulated behind but not in front of the MF. Rdx overexpression (F, marked by GFP in green) reduced levels of Ci155 (left panel, red in right panel) within or just in front of the MF (arrow). (G) When control hsGal4 embryos (+) or hsGal4/MycRdxA embryos (^R) were immunoprecipitated for Myc (IP Myc), Ci155 but not Ci75 was detected in the immunoprecipitates. Neither Ci155 nor Ci75 were detected in control immunoprecipitates using Flag antibody (IP control). Lysates (1% and 0.2% loaded) give estimates of sensitivity. (H) The relationship of Hh, Rdx, Cul3 and Ci. Hh induces Ci155 to activate transcription of rdx (curved red arrow). Then Rdx acts with Cul3 (blue curved arrow) to degrade Ci155 (red blobs). The resulting reduction in Ci155 then attenuates Hh responses (red outlined arrow).