The decoupling of Smoothened from Galphai proteins has little effect on Gli3 protein processing and Hedgehog-regulated chick neural tube patterning

The Hedgehog (Hh) signal is transmitted by two receptor molecules, Patched (Ptc) and Smoothened (Smo). Ptc suppresses Smo activity, while Hh binds Ptc and alleviates the suppression, which results in activation of Hh targets. Smo is a seven-transmembrane protein with a long carboxyl terminal tail. Vertebrate Smo has been previously shown to be coupled to Gα_i proteins, but the biological significance of the coupling in Hh signal transduction is not clear. Here we show that although inhibition of Gα_i protein activity appears to significantly reduce Hh pathway activity in Ptc^−/− mouse embryonic fibroblasts and the NIH3T3-based Shh-light cells, it fails to derepress Shh- or a Smo-agonist-induced inhibition of Gli3 protein processing, a known in vivo indicator of Hh signaling activity. The inhibition of Gα_i protein activity also cannot block the Sonic Hedgehog (Shh)-dependent specification of neural progenitor cells in the neural tube. Consistent with these results, overexpression of a constitutively active Gα_i protein, Gα_i2QL, cannot ectopically specify the neural cell types in the spinal cord, whereas an active Smo, SmoM2, can. Thus, our results indicate that the Smo-induced Gα_i activity plays an insignificant role in the regulation of Gli3 processing and Shh-regulated neural tube patterning.