Identification and characterization of Hedgehog modulator properties after functional coupling of Smoothened to G15.
The seven-transmembrane receptor Smoothened (Smo) transduces the signal initiated by Hedgehog (Hh) morphogen binding to the receptor Patched (Ptc). We have reinvestigated the pharmacological properties of reference molecules acting on the Hh pathway using various Hh responses and a novel functional assay based on the coexpression of Smo with the alpha subunit of the G15 protein in HEK293 cells. The measurement of inositol phosphate (IP) accumulation shows that Smo has constitutive activity, a response blocked by Ptc which indicates a functional Hh receptor complex. Interestingly, the antagonists cyclopamine, Cur61414, and SANT-1 display inverse agonist properties and the agonist SAG has no effect at the Smo-induced IP response, but converts Ptc-mediated inactive forms of Smo into active ones. An oncogenic Smo mutant does not mediate an increase in IP response, presumably reflecting its inability to reach the cell membrane. These studies identify novel properties of molecules displaying potential interest in the treatment of various cancers and brain diseases, and demonstrate that Smo is capable of signaling through G15.
Masdeu
Christelle
C
CNRS, Institut de Neurobiologie Alfred Fessard-IFR 2118, UPR9040, Laboratoire de Neurobiologie Cellulaire et Moléculaire, Signal Transduction and Developmental Neuropharmacology team, 1 avenue de la Terrasse, Gif-sur-Yvette F-91198, France.
Faure
Hélène
H
Coulombe
Josée
J
Schoenfelder
Angèle
A
Mann
André
A
Brabet
Isabelle
I
Pin
Jean-Philippe
JP
Traiffort
Elisabeth
E
Ruat
Martial
M
D000818:Animals; D018797:Cell Cycle Proteins / Q000502:physiology*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004268:DNA-Binding Proteins / Q000502:physiology*; D004149:Dioxoles / Q000494:pharmacology; D016147:Genes, Tumor Suppressor / Q000502:physiology*; D053823:Hedgehog Proteins; D006801:Humans; D051379:Mice; D009369:Neoplasms / Q000378:metabolism; D010879:Piperazines / Q000494:pharmacology; D011720:Pyrazoles / Q000494:pharmacology; D016601:RNA-Binding Proteins / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled / Q000502:physiology*; D015398:Signal Transduction; D000072086:Smoothened Receptor; D013234:Stem Cells / Q000378:metabolism; D015534:Trans-Activators / Q000502:physiology*; D025521:Tumor Suppressor Proteins / Q000502:physiology*; D014704:Veratrum Alkaloids / Q000494:pharmacology
C474318:CUR 61414; D018797:Cell Cycle Proteins; D004268:DNA-Binding Proteins; D004149:Dioxoles; D053823:Hedgehog Proteins; D010879:Piperazines; D011720:Pyrazoles; C473448:RBM5 protein, human; D016601:RNA-Binding Proteins; D043562:Receptors, G-Protein-Coupled; C468919:SANT-1 compound; C491023:SMO protein, human; D000072086:Smoothened Receptor; D015534:Trans-Activators; D025521:Tumor Suppressor Proteins; D014704:Veratrum Alkaloids; C000541:cyclopamine