Assessing Smoothened-mediated Hedgehog signaling in zebrafish

Smoothened belongs to the class of atypical G protein-coupled receptors and serves as the transducing molecule in Hedgehog (Hh) signaling. Hh proteins comprise a family of secreted, cholesterol-modified ligands, which act both as morphogens and as signaling molecules. Binding of Hh proteins to their direct receptor, the transmembrane protein Patched-1, relieves Smoothened from tonal inhibition by Patched-1 and causes the translocation of Smoothened into the cilium. Here, the Hh signaling cascade is initiated and results in transcriptional activation of Hh target genes such as gli1 or patched-1. This induces a plethora of physiological outcomes including normal embryonic development, but also cancer, which is the reason why scientists aim to develop strategies to manipulate as well as monitor Smoothened-mediated Hh signaling. The zebrafish has emerged as a valuable tool for the assessment of Smoothened-mediated Hh signaling. In this chapter we thus describe how Smoothened-mediated Hh signaling can be monitored and also quantified using zebrafish embryos.