UC Berkeley

Sonic Hedgehog (Shh) is a vertebrate signaling molecule that is indispensable for patterning during embryogenesis and for the maintenance of adult stem cell populations. Impaired regulation of the Hedgehog (Hh) pathway can lead to birth defects and disease, thus insight into the regulation of signal transduction is critical for understanding developmental mechanisms and for generating new therapeutic strategies.

The function of the ligand, the receptor, and the signal transducer are separated into distinct molecules and the state of pathway activity depends on their interaction. In the absence of the Shh ligand, the canonical receptors, Patched1/2 (Ptch1/2), are potent inhibitors of Smoothened (Smo), the GPCR-like signal transducer of the Hh pathway. The mechanism by which Shh acts through Ptch1/2 to activate Smo and thereby the Hh response remains unclear.

Here we find that Ptch1LacZ/LacZ;Ptch2-/- cells have a low level of Hh pathway activity and are unresponsive to exogenously supplied ShhN, unless transfected with Ptch1, including forms of Ptch1 lacking antiporter activity required for the inhibition of Smo. However, this Ptch1 requirement can be circumvented by ShhN transfection; Ptch1LacZ/LacZ;Ptch2-/- cells expressing ShhN have an activated Hh response that requires Smo. Mutant forms of Shh that cannot bind Ptch1/2 or the Shh co-receptors, and thus unable to induce the Hh response when applied to Ptch1/2 proficient cells, are nevertheless potent inducers of the Hh response after transfection into Ptch1LacZ/LacZ;Ptch2-/- cells and in vivo. Forms of Smo lacking the N-terminal extracellular Cysteine Rich Domain (CRD) retain their sensitivity to Ptch1-mediated inhibition, but can no longer be activated cell-autonomously by Shh. Our findings support a model in which the role of Ptch1/2 as an allosteric inhibitor of Smo is complemented by a role as a facilitator of a Shh-mediated Smo activation event that is independent of Shh (co-) receptors.