Structural basis of Smoothened regulation by its extracellular domains

Eamon F.X. Byrne, Ria Sircar, Paul S. Miller, George Hedger, Giovanni Luchetti, Sigrid Nachtergaele, Mark D. Tully, Laurel Mydock-Mcgrane, Douglas F. Covey, Robert P. Rambo, Mark S.P. Sansom, Simon Newstead, Rajat Rohatgi, Christian Siebold

Research output: Contribution to journalArticle

156 Scopus citations

Abstract

Developmental signals of the Hedgehog (Hh) and Wnt families are transduced across the membrane by Frizzled-class G-protein-coupled receptors (GPCRs) composed of both a heptahelical transmembrane domain (TMD) and an extracellular cysteine-rich domain (CRD). How the large extracellular domains of GPCRs regulate signalling by the TMD is unknown. We present crystal structures of the Hh signal transducer and oncoprotein Smoothened, a GPCR that contains two distinct ligand-binding sites: one in its TMD and one in the CRD. The CRD is stacked atop the TMD, separated by an intervening wedge-like linker domain. Structure-guided mutations show that the interface between the CRD, linker domain and TMD stabilizes the inactive state of Smoothened. Unexpectedly, we find a cholesterol molecule bound to Smoothened in the CRD binding site. Mutations predicted to prevent cholesterol binding impair the ability of Smoothened to transmit native Hh signals. Binding of a clinically used antagonist, vismodegib, to the TMD induces a conformational change that is propagated to the CRD, resulting in loss of cholesterol from the CRD-linker domain-TMD interface. Our results clarify the structural mechanism by which the activity of a GPCR is controlled by ligand-regulated interactions between its extracellular and transmembrane domains.

Original languageEnglish
Pages (from-to)517-522
Number of pages6
JournalNature
Volume535
Issue number7613
DOIs
StatePublished - Jul 28 2016

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    Byrne, E. F. X., Sircar, R., Miller, P. S., Hedger, G., Luchetti, G., Nachtergaele, S., Tully, M. D., Mydock-Mcgrane, L., Covey, D. F., Rambo, R. P., Sansom, M. S. P., Newstead, S., Rohatgi, R., & Siebold, C. (2016). Structural basis of Smoothened regulation by its extracellular domains. Nature, 535(7613), 517-522. https://doi.org/10.1038/nature18934