Distinct phosphorylations on kinesin costal-2 mediate differential hedgehog signaling strength

The graded Hedgehog (Hh) signal is transduced by the transmembrane Smoothened (Smo) proteins in both vertebrates and invertebrates. In Drosophila, associations between Smo and the Fused (Fu)/Costal-2 (Cos2)/Cubitus Interruptus (Ci) cytoplasmic complex lead to pathway activation, but it remains unclear how the cytoplasmic complex responds to and transduces different levels of Hh signaling. We show here that, within the Hh gradient field, low- and high-magnitude Smo activations control differentially the phosphorylation of Cos2 on two distinct serines. We also provide evidence that these phosphorylations depend on the Fu kinase activity and lead to a shift of Cos2 distribution from the cytoplasm to the plasma membrane. Moreover, the distinct Cos2 phosphorylation states mediate differential Hh signaling magnitude, suggesting that phosphorylation and relocation of Cos2 to the plasma membrane facilitate high-level Hh signaling through the control of Ci nuclear translocation and transcriptional activity.