Mutants of cubitus interruptus that are independent of PKA regulation are independent of hedgehog signaling

Hedgehog (HH) is an important morphogen involved in pattern formation durin g Drosophila embryogenesis and disc development. cubitus interruptus (ci) e ncodes a transcription factor responsible for transducing the hh signal in the nucleus and activating hh target gene expression. Previous studies have shown that CI exists in two forms: a 75 kDa proteolytic repressor form and a 155 kDa activator form. The ratio of these forms, which is regulated pos itively by hh signaling and negatively by PKA activity, determines the on/o ff status of hh target gene expression. In this paper, we demonstrate that the exogenous expression of CI that is mutant for four consensus PKA sites [CI(m1-4)], causes ectopic expression of wingless (wg) in vivo and a phenot ype consistent with wg overexpression. Expression of CI(m1-4), but not CI(w t), can rescue the hh mutant phenotype and restore wg expression in hh muta nt embryos. When PI(A activity is suppressed by expressing a dominant negat ive PKA mutant, the exogenous expression of CP(wt) results in overexpressio n of wg and lethality in embryogenesis, defects that are similar to those c aused by the exogenous expression of CT(m1-4). In addition, we demonstrate that, in cell culture, the mutation of any one of the three serine-containi ng PKA sites abolishes the proteolytic processing of CI, We also show that PKA directly phosphorylates the four consensus phosphorylation sites in vit ro. Taken together, our results suggest that positive hh and negative PI(A regulation of wg gene expression converge on the regulation of CI phosphory lation.