Journal Article

Sterols in an intramolecular channel of Smoothened mediate Hedgehog signaling

Li, Xiaochun, Qi, Xiaofeng, Friedberg, Lucas, Long, Tao, De Bose-Boyd, Ryan

2020

Overview

Smoothened (SMO), a class Frizzled G protein-coupled receptor (class F GPCR), transduces the Hedgehog signal across the cell membrane. Sterols can bind to its extracellular cysteine-rich domain (CRD) and to several sites in the seven transmembrane helices (7-TMs) of SMO. However, the mechanism by which sterols regulate SMO via multiple sites is unknown. Here we determined the structures of SMO–G i complexes bound to the synthetic SMO agonist (SAG) and to 24( S ),25-epoxycholesterol (24( S ),25-EC). A novel sterol-binding site in the extracellular extension of TM6 was revealed to connect other sites in 7-TMs and CRD, forming an intramolecular sterol channel from the middle side of 7-TMs to CRD. Additional structures of two gain-of-function variants, SMO D384R and SMO G111C/I496C , showed that blocking the channel at its midpoints allows sterols to occupy the binding sites in 7-TMs, thereby activating SMO. These data indicate that sterol transport through the core of SMO is a major regulator of SMO-mediated signaling. Cryo-EM structural work shows sterols binding at four adjacent locations within the class F GPCR Smoothened (SMO), where the transmembrane core functions as a sterol tunnel in which occupancy activates SMO for downstream Hedgehog signaling.

• Publisher: Nature Publishing Group US,Nature Publishing Group
• Subject: 631/45/287/1197 / 631/535 / 631/80/86 / 631/92/612/194 / Amino Acid Sequence / Amino Acid Substitution / Binding Sites / Biochemical Engineering / Biochemistry / Bioorganic Chemistry / Cell Biology / Cell membranes / Chemistry / Chemistry and Materials Science / Chemistry/Food Science / Cholesterol - analogs & derivatives / Cholesterol - chemistry / Cholesterol - metabolism / Cyclohexylamines - chemistry / Cyclohexylamines - pharmacology / Frizzled protein / G protein-coupled receptors / Gene Expression Regulation / GTP-Binding Protein alpha Subunits - chemistry / GTP-Binding Protein alpha Subunits - genetics / GTP-Binding Protein alpha Subunits - metabolism / GTP-Binding Protein beta Subunits - chemistry / GTP-Binding Protein beta Subunits - genetics / GTP-Binding Protein beta Subunits - metabolism / GTP-Binding Protein gamma Subunits - chemistry / GTP-Binding Protein gamma Subunits - genetics / GTP-Binding Protein gamma Subunits - metabolism / Hedgehog protein / HEK293 Cells / Helices / Humans / Molecular Docking Simulation / Occupancy / Protein Binding / Protein Conformation, alpha-Helical / Protein Conformation, beta-Strand / Protein Interaction Domains and Motifs / Recombinant Proteins - chemistry / Recombinant Proteins - genetics / Recombinant Proteins - metabolism / Sequence Alignment / Sequence Homology, Amino Acid / Signal Transduction / Signaling / Single-Chain Antibodies / Smoothened Receptor - agonists / Smoothened Receptor - chemistry / Smoothened Receptor - genetics / Smoothened Receptor - metabolism / Sterols / Thiophenes - chemistry / Thiophenes - pharmacology