Hedgehog Signalling

Abstract

Hedgehog (Hh) proteins are a family of secreted factors with morphogen activities that have essential role during embryonic development and in adulthood. Mechanisms underlying the production and reception, as well as the signal transduction pathway of Hh proteins, are highly conserved between invertebrates and vertebrates. However, despite this evolutionary conservation, a major divergence is the pathway's association with primary cilia in vertebrates. Hh proteins act both as short‐range and long‐range factors in the control of cell fate specification and cell differentiation, cell proliferation, tissue patterning and morphogenesis during embryonic development. Hh proteins are also involved in tumour formation. Knowledge of this pathway has been instrumental in recent therapeutic approaches aiming at downregulating Hh signalling in cancers.

Key Concepts:

  • Hedgehog signalling is conserved during evolution.

  • Hedgehog is a morphogen, specifying distinct cell types at different concentrations.

  • Hedgehog acts as a short‐ and long‐range signalling molecule.

  • Hedgehog has essential role in cell fate specification and differentiation and in cell proliferation and survival.

  • Hedgehog signalling is implicated in cancer.

Keywords: embryonic development; cell signalling; Hedgehog; morphogen; cell fate specification; cell differentiation; cell proliferation; cancer

Figure 1.

Synthesis and release of Hh proteins. Hh proteins are synthesised as precursors that undergo autocatalytic cleavage and lipid modifications in the endoplasmic reticulum. These modifications include cholesterol incorporation to facilitate anchoring to the plasma membrane and Ski/HHAT‐mediated palmitic acid addition to enhance secretion. Hh proteins can be released via Dispatched‐mediated action and the cooperative binding of Scube protein. Other release mechanisms involve the recruitment of Hh‐N multimers within lipid vesicles that are coupled to Glypican and Lipophorin (Lipoproteins in vertebrates) complexes.

Figure 2.

Hh signal transduction pathway. (a) Signalling pathway in Drosophila. In the absence of Hh ligand (inactive), Ptc blocks the activity of Smo and promotes its degradation. Here the motor protein Cos2 associates with the HSC components, including the kinases PKA, CKI, GSK3β, Fu and Ci, and binds to microtubules. The HSC complex promotes the sequential phosphorylation of full‐length Ci to generate a repressor (CiR). On Hh binding to Ptc (active), Smo translocates to the plasma membrane, becomes phosphorylated and binds Cos2. Subsequently, Fu‐mediated phosphorylation of Cos2 and Su(Fu) causes the dissociation of the HSC and the release of full‐length Ci, which translocates to the nucleus to function as a transcriptional activator (CiA). (b) Signalling pathway in vertebrates. In vertebrates, Hh signalling transduction is associated with primary cilia. In the absence of Hh, Ptch1 accumulates in primary cilia and prevents Smo activity. Kif7 (Cos2 homologue) accumulation at the basis of cilia promotes Gli2 and Gli3 enrichment at the basis of cilia, where they are phosphorylated by PKA, CKI and GSK3β, and proteolytically cleaved to generate a repressor form (GliR). Hh binding to Ptch1 is potentiated by coreceptors Cdo and Boc and by Gas1. This event promotes Ptch1 trafficking out of the primary cilium and the accumulation of Smo in the axonema, where it is phosphorylated. Kif7 (Cos2 homologue) translocates to the tip of the cilia, allowing the accumulation of Gli2 and Gli3 at the cilia tip and the dissociation of the SuFu–Gli complex. Full‐length Gli proteins are converted into Gli activator proteins (GliA) and transported out of the cilia to the nucleus.

Figure 3.

Short‐range Hh signalling. (a) In the Drosophila epidermis, restricted expression domains of Wingless and Hedgehog proteins along the A–P axis patterns the fly cuticle. wg is expressed in anterior cells and activates engrailed (en) expression in posterior cells, which, in turn, induces hh expression. Subsequently, both wg and hh restrict the expression of serrate (ser) within two to three cells per PS. Next, both hh and ser induce rhomboid (rho) expression to generate a three‐cell‐wide stripe anteriorly adjacent to the ser expression domain. Wg also blocks the expression of rho in anterior cells. The mutually exclusive expression domains of ser and rho will dictate epidermal cell fate and eventually the formation of spiked and smooth cuticle. (b) In the fly wing imaginal disc, posterior cells produced hh (blue domain), which acts as a morphogen in cells within the A/P boundary and the anterior compartment. There Hh induces the expression of specific genes, such as en, ptc, col and dpp to form different expression gradients responsible for wing patterning. Although high levels of Hh are required to induce ptc, en and col expression, dpp responds to lower Hh levels.

Figure 4.

Long‐range Hh signalling patterns the spinal cord. Shh produced by the notochord and the floor plate (blue dots) creates a gradient of at least 15–20‐cell diameter along the D‐V axis, with high Shh concentration in the ventral domain and low Shh concentration in the central domain of the neural tube. At early stages, this gradient induces the specification of distinct progenitor cells (p3 to p0) according to the threshold of Shh cells exposed along the D–V axis of the spinal cord. These thresholds translate into a gradient of Gli activity, with Gli3R inhibited by low levels of Shh in ventral domains (pMN, p2 and p1) and Gli2 and Gli1 acting as activators at high levels of Shh in the most ventral domains (p3 and pMN). Each progenitor domain expresses a particular combination of transcription factors (Pax7, FoxA2, Pax6, Irx3, Dbx1, Nkx2.2, Olig2, Dbx2, Nkx6.1 and others), which are also regulated by Shh in a dose‐dependent manner. Boundaries of progenitor domains are subsequently reinforced through mutual negative regulation between markers of adjacent domains, and progenitor cells within each domain differentiate into specific populations of neurons (V3 to V0).

Figure 5.

Temporal control of the Hh signalling pathway. (a) Patterning and specification of the vertebrate limb is subject to Shh control produced by the ZPA (blue area), which creates a gradient along the A–P axis. During early limb bud development, Shh secretion overcomes Gli3R inhibition in the posterior limb, inducing an opposite gradient of Gli3R activity. At later stages, Shh controls digit specification in a time‐ and dose‐dependent fashion. Here although the most posterior digits (5 and 4 in mice) are exposed to the highest morphogen levels for longer periods of time, intermediate and anterior digits are exposed to lower Shh levels. (b) Shh produced by the neural tube and the notochord (blue dots) controls muscle fibre specification in a dose‐ and temporal‐dependent manner in the zebrafish embryo. Adaxial cells, which are adjacent to the notochord, contain precursor cells for MP cells (red) and for SSF (green). MPs differentiate in situ, whereas SSFs migrate to the surface of somites. MFFs (orange) and fast fibres (white) differentiate behind migrating SSFs. Higher levels of Shh are required to specify MPs, compared with SSFs. MFFs are specified by exposure to intermediate levels of Shh, whereas fast fibres do not depend on Shh signalling. In addition, MPs and MFFs required longer exposure to Shh than SSFs. Muscle fibre specification can also be explained by a 3D histogram representing the relationship between Shh levels and exposure time to Shh required to generate distinct muscle fibre types. Here MP cells (red bar) are exposed to high levels of Shh for a longer period of time than MFFs (orange bar) and SFFs (green bar). Similarly, MFFs require longer exposure to intermediate levels of Shh than SFFs, which are specified by low levels of Shh for a short period of time.

Figure 6.

Hedgehog acts as a mitogen in the cerebellum and retina. (a) In the cerebellum, Shh secreted by Purkinje cells positively controls the proliferation of cerebellar granular neural precursors (blue circles) within the external granular layer. Specifically, Shh induces the expression of D‐type cyclins and other proteins, such as Nmyc1 via Gli1 and Gli2 activity. This regulation impacts on the size and foliation of the cerebellum postnatally. (b) During early retinogenesis of lower vertebrates, locally secreted Shh (blue dots) induces the proliferation of RPCs in the ciliary marginal zone (1) by controlling the expression of G1 cyclins, such as cyclin D1. However, at later time points, Shh appears to control cell cycle through the regulation of G2–M transition regulators, such as Cyclin B1 and Cdc25c. Thus, Shh has a dual function in RPC development: first promoting the expansion of RPCs and then subsequently acting negatively on retinal ganglion progenitor cells by promoting cell cycle exit and differentiation (2).

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Further Reading

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Varjosalo M and Taipale J (2008) Hedgehog: functions and mechanisms. Genes & Development 22: 2454–2472.

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Cruz‐Migoni, Sara Betania, and Borycki, Anne‐Gaëlle(Mar 2014) Hedgehog Signalling. In: eLS. John Wiley & Sons Ltd, Chichester. http://www.els.net [doi: 10.1002/9780470015902.a0000806.pub2]