Hedgehog signaling through the ciliary membrane protein Smoothened dissociates the Gli transcription factors from their inhibitor, Suppressor of Fused.

Summary The Hedgehog signaling pathway plays critical roles in metazoan development and in cancer. How the Hedgehog ligand is secreted and spreads to distant cells is unclear, given its covalent modification with a hydrophobic cholesterol molecule, which makes it stick to membranes. We demonstrate that Hedgehog ligand secretion from vertebrate cells is accomplished via two distinct and synergistic cholesterol-dependent binding events, one mediated by the membrane protein Dispatched and the other by a Scube family member, secreted proteins essential for Hedgehog signaling in zebrafish. Cholesterol modification is sufficient for a heterologous protein to interact with Scube, and to be secreted in a Scube-dependent manner. Dispatched and Scube recognize different structural aspects of cholesterol, similar to how Niemann-Pick disease proteins 1 and 2 interact with cholesterol, suggesting a hand-off mechanism for transferring Hedgehog from Dispatched to Scube. Thus, Dispatched and Scube cooperate to dramatically enhance secretion and solubility of the cholesterol-modified Hedgehog ligand.

In higher eukaryotes, the endoplasmic reticulum (ER) contains a network of membrane tubules, which transitions into sheets during mitosis. Network formation involves curvature-stabilizing proteins, including the reticulons (Rtns), as well as the membrane-fusing GTPase atlastin (ATL) and the lunapark protein (Lnp). Here, we have analyzed how these proteins cooperate. ATL is needed to not only form, but also maintain, the ER network. Maintenance requires a balance between ATL and Rtn, as too little ATL activity or too high Rtn4a concentrations cause ER fragmentation. Lnp only affects the abundance of three-way junctions and tubules. We suggest a model in which ATL-mediated fusion counteracts the instability of free tubule ends. ATL tethers and fuses tubules stabilized by the Rtns, and transiently sits in newly formed three-way junctions. Lnp subsequently moves into the junctional sheets and forms oligomers. Lnp is inactivated by mitotic phosphorylation, which contributes to the tubule-to-sheet conversion of the ER.DOI: http://dx.doi.org/10.7554/eLife.18605.001