Fig. 2. Specific immunodetection of the Hh-induced phosphorylated form of Cos2 on Ser572. (A) On western blot analysis, the anti-S572P antibody specifically detects the Hh-induced phosphorylated form of Cos2 on Ser572. Notice the absence of signal in the non-Hh-expressing cells and the strong decrease in the Hh-expressing cells treated with fu RNAi (lane 2 in right panel). (B) After transfection with cos2 and fu constructs, S2 cells were treated with okadaic acid (OA) to mimic Hh pathway activation. Notice that phosphorylated Cos2 was absent when cells were transfected with Fu-KD. (C) S2 cells were transfected with cos2-WT-myc, fu-WT-HA and different variants of smo. Phosphorylated Cos2 on Ser572 was increased in the presence of the SmoDDD variant but was absent in the presence of SmoAAA or Fu-KD.