Hedgehog signalling in gut development, physiology and cancer

Abstract

The Hedgehog pathway is one of the most common signal transduction pathways used by mammalian cells. Most studies have focused on its role during development, primarily of the nervous system, skin, bone and pancreas. Due to the activation of this pathway during proliferation and neoplastic transformation, more recent studies have examined its role in adult tissues. Significant levels of sonic hedgehog are expressed in the gastric mucosa, which has served to direct analysis of its role during organogenesis, gastric acid secretion and neoplastic transformation. Therefore the goal of this review is to apply current knowledge of this pathway to further our understanding of gastrointestinal physiology and neoplasia, using the stomach as a prototype.

Figure 1. Sonic Hedgehog processing

Two mechanisms of Shh processing have been reported. The best known is the autocatalytic mechanism of Shh processing in which the C-terminus functions as a cholesterol esterase adding the sterol to cysteine 199 followed by palmitoylation at residue 25. The fatty acid permits Shh to be tethered to the plasma membrane until it is cleaved by an ADAM protease. The cleaved Shh molecules form miscelles in the presence of the protein Dispatched (Disp). This mechanism has been described for Drosophila cells and cells derived from mesenchyme. By contrast, gastric acid producing parietal cells cleave Shh in a protease (pepsin)-dependent manner. It is not known whether Shh produced from the parietal cell is post-translationally modified with cholesterol or a fatty acid.

Figure 2. Schematic diagram of Gli transcription factors

Gli1, Gli2 and Gli3 are zinc finger homologues of Drosophila Cubitus interruptus (Ci). The 4 transcription factors are aligned to demonstrate the homology between the various isoforms. Various functional domains of these proteins are indicated. Znf, zinc finger domain; TAD, transcription activation domain; Rep, repressor domain; SuFu BS, suppressor of fused binding site; PC, phosphorylation cluster; *protein kinase A phosphorylation site; cleavage domain: site of cleavage by proteasome. Modified from Hui & Angers, 2011 with permission.

Figure 3. Hedgehog signalling in primary cilia

A, in the absence of Shh ligand, Smo-laden vesicles reside in the cytoplasm. Full-length (FL) Gli2 and Gli3 reside at the cilia tip with repressor complex proteins Fused, SuFu and Kif7. The intraflagellar protein (IFT) moves into the cilium with cargo proteins. If Gli2 and 3 are phosphorylated sequentially by PKA, CK or GSK3β, the docking protein βTrCP recognizes the phosphopeptide motif and recruits an E3 ligase to ubiquitinate (Ub) the protein. Ub Gli proteins undergo degradation by the proteasome. The repressor Gli2 protein moves to the nucleus and inhibits genes with co-repressor complexes. B, Hh signalling triggers Smo-laden vesicles to fuse with the plasma membrane once relieved from the inhibitory effects of the receptor Ptch. IFTs carry Smo antegrade to the tip of the cilium. Fused phosphorylates SuFu, Gli2 and Gli3 are released to the cytoplasm, phosphorylation is inhibited and Gli3 repressor is degraded. Gli2 migrates to the nucleus to bind Hh target genes.

Figure 4. Gastric Shh inversely correlated with mucous cell compartment expansion

A significant amount of Shh in the stomach is generated by the acid-producing parietal cells. Conditionally null Shh mice expand their surface pit mucous cell compartment (foveolar hyperplasia). Shh processing in stomach requires acid secretion and is stimulated by gastrin. Shh is known to form a concentration gradient and in turn exert a differential effect on genes as a result. Thus there is a sliding scale of Shh concentrations and subsequent effects on gene expression. Therefore gastric Shh might form a gradient with increasing Shh concentration towards the gastric gland lumen (scenario no. 1) or maintain a gradient in which the concentrations are elevated near the mid-gastric gland then diffuse toward the lumen and gland base (scenario no. 2). Examples of potential gene targets expressed by specific cell types are listed on the right.

Figure 5. Pro-inflammatory cytokine initiation of Shh feedforward loop

During chronic gastric inflammation, pro-inflammatory cytokines (Th1) are able to stimulate Shh gene expression in stromal cells. Inflammation in the stomach inhibits parietal cell acid secretion and eventually triggers loss of the parietal and zymogenic cell lineages (atrophy). This potentially results in expansion of the mesenchymal cell population and production of pro-proliferative stromal factors, e.g. Wnts, TGFβ. Wnt signalling is then available to induce mucous cell proliferation (metaplasia) and subsequently dysplasia/cancer.