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Sonic Hedgehog

Template:STRUCTURE 1vhh

Introduction

Sonic hedgehog (Shh) is a member of the Hedgehog (Hh) family of secreted extracellular signaling proteins, which serve important roles in regulating both short-range and long-range patterning processes in developing invertebrate and vertebrate tissues ^[1]. First discovered in Drosophila, where mutations of the single Hedgehog gene produces larvae that are covered in hedgehog-like denticles, Hh proteins are encoded by at least three genes in mammals - Sonic, Desert, and Indian hedgehog ^[2]. With the ability to control such fundamental processes as pattern formation in vertebrate limb buds ^[3], the formation of motor neurons in the neural tube ^[4], and the development and maintenance of tissues and organs, Shh is the most well-studied member of the Hh signaling pathway. Excessive signaling in adult cells has been implicated in the development of several human cancers ^[5][6].

Biosynthesis

As with all members of the Hh family, Shh biosynthesis begins with an unusual molecular processing event. Following cleavage of its signal peptide, the Shh precursor protein is autocatalytically cleaved into two functionally distinct domains, a 19-kDa amino-terminal domain (Shh-N) and a 27-kDa carboxy-terminal domain (Shh-C) ^[7]. Spanning residues 24 to 197 in human Shh, Shh-N is responsible for all of the local and long-range signaling activities of Shh. Shh-C possesses an intramolecular transferase activity responsible for covalent attachment of a molecule of cholesterol to the C-terminus of Shh-N. The addition of cholesterol serves to tether Shh-N to the cell membrane, restricting its range of activity to that of local signaling only ^[8]. A second modification involving the attachment of a palmitoyl group to Cys-24 on the protein's N-terminus has recently been discovered in insect and mammalian cells. This N-terminal modification is thought to increase the potency of the Shh-N signal as much as 30-fold ^[9].

Structural Overview

An α + β sandwich consisting of two and a six-stranded, mixed makes up the core of the native Shh-N structure, along with a two-stranded, antiparallel β-sheet. Although this type of folding arrangement has not yet been seen in other proteins, the presence of a in Shh-N bears close structural resemblance to the zinc coordination sites of zinc hydrolases, including thermolysin and carboxypeptidase A. Three amino acid side chains – – are bound to the zinc ion in the crystal structure, along with a single . Zinc ions that serve a structural role in proteins are normally coordinated by four amino acid side chains, including a cysteine, and are not usually exposed to the surrounding solvent. The presence of a zinc-bound water molecule in Shh-N, by contrast, is indicative of a catalytic function. In zinc hydrolases, the water molecule is key to the protein's enzymatic activity when its proton is removed by a nearby glutamate residue. likely serves the same role in Shh-N, further supporting a novel, hydrolytic function for this protein. Based on the catalytic mechanisms for thermolysin and carboxypeptidase A, in Shh-N are also believed to participate in a potential hydrolysis reaction ^[10].

Function

The tetrahedrally coordinated zinc ion of Shh-N, along with the non-coordinating residues thought to assist hydrolysis, are highly conserved among vertebrate Hh proteins. A potential hydrolytic activity is therefore expected to play an important cellular role. In pursuit of a substrate for Shh-N, it was found that Ala 194 and Lys 195 near the C-terminus of one Shh-N molecule can hydrogen bond with residues in the zinc binding site of a second Shh-N molecule. This indicates that the protein may be capable of cleaving its own C-terminus, a function that has been suggested to liberate the tethered Shh-N from the cell membrane to facilitate long-range signaling ^[10]

Sonic Signaling: The Shh-Gli Pathway

Sonic Hedgehog signaling pathway. In the absence of Shh, Patched inhibits Smo. Inhibition of Patched by Shh activates normal developmental processes. [Note: This figure is adapted from references ^[6] and ^[11].]

In the absence of a Shh signal, a transmembrane receptor protein called Patched blocks the function of Smoothened (Smo), a seven-pass transmembrane protein, by keeping it sequestered in an intracellular vesicle. When Shh binds to Patched, inhibition of Smo by Patched is relieved. Patched becomes endocytosed, and Smo translocates to the cell surface. In vertebrates, Smo localizes to the surface of the primary cilium, initiating a signaling cascade that leads to the activation of Gli transcription factors ^[6]. Present in both the nucleus and cytoplasm, there are three of these regulatory proteins (Gli1, Gli2, and Gli3). Following Shh signaling, all three proteins can act as transcriptional activators of Shh target genes. Gli3, however, can act as both an activator and repressor; in the absence of Shh signaling, Gli3 is cleaved by the proteasome, and its truncated form accumulates in the nucleus where it represses transcription of Shh-responsive genes ^[11].

Mutations

References

1. ↑ Perrimon N. Hedgehog and beyond. Cell. 1995 Feb 24;80(4):517-20. PMID:7867057
2. ↑ Echelard Y, Epstein DJ, St-Jacques B, Shen L, Mohler J, McMahon JA, McMahon AP. Sonic hedgehog, a member of a family of putative signaling molecules, is implicated in the regulation of CNS polarity. Cell. 1993 Dec 31;75(7):1417-30. PMID:7916661
3. ↑ Riddle RD, Johnson RL, Laufer E, Tabin C. Sonic hedgehog mediates the polarizing activity of the ZPA. Cell. 1993 Dec 31;75(7):1401-16. PMID:8269518
4. ↑ Goodrich LV, Scott MP. Hedgehog and patched in neural development and disease. Neuron. 1998 Dec;21(6):1243-57. PMID:9883719
5. ↑ Pasca di Magliano M, Hebrok M. Hedgehog signalling in cancer formation and maintenance. Nat Rev Cancer. 2003 Dec;3(12):903-11. PMID:14737121 doi:10.1038/nrc1229
6. ↑ ^{6.0 6.1 6.2} Ruiz i Altaba A, Sanchez P, Dahmane N. Gli and hedgehog in cancer: tumours, embryos and stem cells. Nat Rev Cancer. 2002 May;2(5):361-72. PMID:12044012 doi:10.1038/nrc796
7. ↑ Bumcrot DA, Takada R, McMahon AP. Proteolytic processing yields two secreted forms of sonic hedgehog. Mol Cell Biol. 1995 Apr;15(4):2294-303. PMID:7891723
8. ↑ Porter JA, Young KE, Beachy PA. Cholesterol modification of hedgehog signaling proteins in animal development. Science. 1996 Oct 11;274(5285):255-9. PMID:8824192
9. ↑ Pepinsky RB, Zeng C, Wen D, Rayhorn P, Baker DP, Williams KP, Bixler SA, Ambrose CM, Garber EA, Miatkowski K, Taylor FR, Wang EA, Galdes A. Identification of a palmitic acid-modified form of human Sonic hedgehog. J Biol Chem. 1998 May 29;273(22):14037-45. PMID:9593755
10. ↑ ^{10.0 10.1} Hall TM, Porter JA, Beachy PA, Leahy DJ. A potential catalytic site revealed by the 1.7-A crystal structure of the amino-terminal signalling domain of Sonic hedgehog. Nature. 1995 Nov 9;378(6553):212-6. PMID:7477329 doi:http://dx.doi.org/10.1038/378212a0
11. ↑ ^{11.0 11.1} Huangfu D, Anderson KV. Signaling from Smo to Ci/Gli: conservation and divergence of Hedgehog pathways from Drosophila to vertebrates. Development. 2006 Jan;133(1):3-14. PMID:16339192 doi:133/1/3