Hh-induced Smoothened conformational switch is mediated by differential phosphorylation at its C-terminal tail in a dose- and position-dependent manner

Dev Biol. 2012 Jun 15;366(2):172-84. doi: 10.1016/j.ydbio.2012.04.007. Epub 2012 Apr 19.


The activation of Smoothened (Smo) requires phosphorylation at three clusters of Serine residues in Drosophila Hedgehog (Hh) signaling. However, the mechanism by which phosphorylation promotes Smo conformational change and subsequently activates Smo in response to Hh gradient remains unclear. Here, we show that the conformational states of Smo are determined by not only the amount but also the position of the negative charges provided by phosphorylation. By using a Smo phospho-specific antibody, we demonstrate that Smo is differentially phosphorylated at three clusters of serine residues in response to levels of Hh activity. Mutating the first cluster, compared to mutating the other clusters, impairs Smo activity more severely, whereas mutating the last cluster prohibits C-terminus dimerization. In addition, phosphorylation of the membrane proximal cluster promotes phosphorylation of the distal cluster. We propose a zipper-lock model in which the gradual phosphorylation at these clusters induces a gradual conformational change in the Smo cytoplasmic tail, which promotes the interaction between Smo and Costal2 (Cos2). Moreover, we show that Hh regulates both PKA and CK1 phosphorylation of Smo. Thus, the differential phosphorylation of Smo mediates the thresholds of Hh activity.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Drosophila / metabolism*
  • Drosophila Proteins / chemistry
  • Drosophila Proteins / metabolism*
  • Hedgehog Proteins / metabolism*
  • Models, Molecular
  • Phosphorylation
  • Protein Conformation
  • Protein Structure, Tertiary
  • Receptors, G-Protein-Coupled / chemistry
  • Receptors, G-Protein-Coupled / metabolism*
  • Signal Transduction
  • Smoothened Receptor


  • Drosophila Proteins
  • Hedgehog Proteins
  • Receptors, G-Protein-Coupled
  • Smoothened Receptor
  • smo protein, Drosophila