The sonic hedgehog receptor patched associates with caveolin-1 in cholesterol-rich microdomains of the plasma membrane

The hedgehog signaling pathway is involved in early embryonic patterning as well as in cancer; however, little is known about the subcellular localiza tion of the Hedgehog receptor complex of Patched and Smoothened. Since Hh h as been found in lipid rafts in Drosophila, we hypothesized that Patched an d Smoothened might also be found in these cholesterol-rich microdomains. In this study, we demonstrate that both Smoothened and Patched are in caveoli n-1-enriched/raft microdomains. Immunoprecipitation studies show that Patch ed specifically interacts with caveolin-1, whereas Smoothened does not. Fra ctionation studies show that Patched and caveolin-1 can be co-isolated from buoyant density fractions that represent caveolae/raft microdomains and th at Patched and caveolin-1 co-localize by confocal microscopy, Glutathione S -transferase fusion protein experiments show that the interaction between P atched and caveolin-1 involves the caveolin-1 scaffolding domain and a Patc hed consensus binding site. Immunocytochemistry data and fractionation stud ies also show that Patched seems to be required for transport of Smoothened to the membrane. Depletion of plasmalemmal cholesterol influences the dist ribution of the Hh receptor complex in the caveolin-enriched/raft microdoma ins. These data suggest that caveolin-1 may be integral for sequestering th e Hh receptor complex in these caveolin-enriched microdomains, which act as a scaffold for the interactions with the Hh protein.