The effects of Hedgehog are mediated by a latent transcription regulator called Cubitus interruptus (Ci), the regulation of which is reminiscent of the regulation of β-catenin by Wnts. In the absence of a Hedgehog signal, Ci is ubiquitylated and proteolytically cleaved in proteasomes. Instead of being completely degraded, however, Ci is processed to form a smaller fragment, which accumulates in the nucleus, where it acts as a transcriptional repressor, helping to keep Hedgehog-responsive genes silent. The proteolytic processing of the Ci protein depends on its phosphorylation by three protein kinases—PKA and two kinases also used in the Wnt pathway, namely GSK3 and CK1. As in the Wnt pathway, the proteolytic processing occurs in a multiprotein complex. The complex includes the protein kinase Fused and a scaffold protein Costal2, which stably associates with Ci, recruits the three other kinases, and binds the complex to microtubules, thereby keeping unprocessed Ci out of the nucleus (Figure 15–61A).
Patched inactive Smoothened microtubule Costal2 Fused large Ci protein PHOSPHORYLATION OF Ci BY PKA, GSK3, AND CK1 iHog Ig-like domain fbronectin-type-III-like domain vesicle cleaved Ci protein in complex with co-repressor
Hedgehog functions by blocking the proteolytic processing of Ci, thereby changing it into a transcriptional activator. It does this by a convoluted signaling process that depends on three transmembrane proteins: Patched, iHog, and Smoothened. Patched is predicted to cross the plasma membrane 12 times, and, although much of it is in intracellular vesicles, some is on the cell surface where it can bind the Hedgehog protein. iHog is also on the cell surface and is thought to serve as a co-receptor for Hedgehog. Smoothened is a seven-pass transmembrane protein with a structure very similar to a GPCR, but it does not seem to act as a Hedgehog receptor or even as an activator of G proteins; it is controlled by Patched and iHog.
In the absence of a Hedgehog signal, Patched employs an unknown mechanism to keep Smoothened sequestered and inactive in intracellular vesicles (see Figure 15–61A). The binding of Hedgehog to iHog and Patched inhibits the activity of Patched and induces its endocytosis and degradation. The result is that Smoothened is liberated from inhibition and translocates to the plasma membrane, where it recruits the protein complex containing Ci, Fused, and Costal2.
Figure 15–61 Hedgehog signaling in Drosophila. (A) In the absence of Hedgehog, most Patched is in intracellular vesicles (not shown), where it keeps Smoothened inactive and sequestered. The Ci protein is bound in a cytosolic protein degradation complex, which includes the protein kinase Fused and the scaffold protein Costal2. Costal2 recruits three other protein kinases (PKA, GSK3, and CK1; not shown), which phosphorylate Ci. Phosphorylated Ci is ubiquitylated and then cleaved in proteasomes (not shown) to form a transcriptional repressor, which accumulates in the nucleus to help keep Hedgehog target genes inactive.
(B) Hedgehog binding to iHog and Patched removes the inhibition of Smoothened by Patched. Smoothened is phosphorylated by PKA and CK1 and translocates to the plasma membrane, where it recruits the complex containing Fused, Costal2, and Ci. Costal2 releases unprocessed Ci, which accumulates in the nucleus and activates the transcription of Hedgehog target genes. Many details in the pathway are poorly understood, including the role of Fused.