|Alkaline Phosphatase Production Induced by Indian Hedgehog/Ihh and Neutralization by Mouse Indian Hedgehog/Ihh Antibody. Recombinant Human/Mouse Indian Hedgehog/Ihh (C28II), N-Terminus induces alkaline phosphatase production in the C3H10T1/2 mouse embryonic fibroblast cell line in a dose-dependent manner (orange line). Alkaline phosphatase production elicited by Indian Hedgehog/Ihh (10 ug/mL) is neutralized (green line) by increasing concentrations of Rat Anti-Mouse Indian Hedgehog/Ihh Monoclonal Antibody (Catalog # MAB8048). The ND50 is typically 10-45 ug/mL.|
The hedgehog (hh) gene encoding a secreted protein was originally identified in Drosophila as a segment polarity gene. The vertebrate homologues of Hh comprise several proteins including Sonic hedgehog (Shh), Indian hedgehog (Ihh), and Desert hedgehog (Dhh) (1). Hedgehog proteins are important signaling molecules during embryonic development and are highly conserved within and across species (1). Mouse and human Ihh share 100% amino acid (aa) identity in the signaling domain, while mouse Ihh and Shh share 90% aa identity in the N-terminal signaling domain. Ihh mRNA expression is detected in fetal lung, gut, stomach, liver, kidney, pancreas and strongly in cartilage, in growth regions of the developing bone (2, 3). Ihh, along with parathyroid hormone related protein, regulate the rate of chondrocyte proliferation and differentiation (4). Ihh is also involved in yolk sac vasculogenesis, playing an important role in differentiation of epiblast cells into endothelial and red blood cells (5). Mouse Ihh cDNA encodes a 411 aa polypeptide with a predicted 27 aa signal peptide. This polypeptide is cleaved to generate a 45 kDa precursor protein that undergoes the same post-translation processing as Shh (3). An autocatalytic reaction yields a 19 kDa amino-terminal domain Ihh-N protein that retains all known signaling capabilities, and a 23 kDa carboxy-terminal domain Ihh-C protein (3). Since hydrophobic modifications to Shh, including the substitution of the N-terminal cysteine residue with two hydrophobic isoleucine residues, can also increase its potency (6), a similar modification was made for Ihh. This modified form also shows increased potency in a bioassay measuring induction of alkaline phosphatase. At the cell surface, Hedgehog activity is mediated by a multicomponent receptor complex involving the 12-pass transmembrane protein Patched (Ptc) which binds Hedgehogs with high affinity and Smoothened (Smo), a signaling seven transmembrane G-protein coupled receptor (1).