Sonic hedgehog induces the segregation of patched and smoothened in endosomes.
BACKGROUND
Sonic hedgehog (Shh) signal transduction involves the ligand binding Patched1 (Ptc1) protein and a signaling component, Smoothened (Smo). A select group of compounds inhibits both Shh signaling, regulated by Ptc1, and late endosomal lipid sorting, regulated by the Ptc-related Niemann-Pick C1 (NPC1) protein. This suggests that Ptc1 regulates Smo activity through a common late endosomal sorting pathway also utilized by NPC1. During signaling, Ptc accumulates in endosomal compartments, but it is unclear if Smo follows Ptc into the endocytic pathway.


RESULTS
We characterized the dynamic subcellular distributions of Ptc1, Smo, and activated Smo mutants individually and in combination. Ptc1 and Smo colocalize extensively in the absence of ligand and are internalized together after ligand binding, but Smo becomes segregated from Ptc1/Shh complexes destined for lysosomal degradation. In contrast, activated Smo mutants do not colocalize with nor are cotransported with Ptc1. Agents that block late endosomal transport and protein sorting inhibit the ligand-induced segregation of Ptc1 and Smo. We show that, like NPC1-regulated lipid sorting, Shh signal transduction is blocked by antibodies that specifically disrupt the internal membranes of late endosomes, which provide a platform for protein and lipid sorting.


CONCLUSIONS
These data support a model in which Ptc1 inhibits Smo only when in the same compartment. Ligand-induced segregation allows Smo to signal independently of Ptc1 after becoming sorted from Ptc1/Shh complexes in the late endocytic pathway.
Department of Biological Structure and Center for Developmental Biology, University of Washington, Seattle 98195, USA.
John P
JP
Incardona
Jean
J
Gruenberg
Henk
H
Roelink
D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D004330:Drosophila; D029721:Drosophila Proteins; D011992:Endosomes / Q000502:physiology*; D053823:Hedgehog Proteins; D008565:Membrane Proteins / Q000502:physiology*; D000072079:Patched Receptors; D011956:Receptors, Cell Surface / Q000502:physiology*; D043562:Receptors, G-Protein-Coupled; D015398:Signal Transduction; D000072086:Smoothened Receptor; D015534:Trans-Activators / Q000502:physiology*; D014714:Vertebrates
D029721:Drosophila Proteins; D053823:Hedgehog Proteins; D008565:Membrane Proteins; D000072079:Patched Receptors; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D000072086:Smoothened Receptor; D015534:Trans-Activators; C100664:smo protein, Drosophila