The Dispatched protein, which is related to the NPC1 and PTCH1 cholesterol transporters^1,2 and to H⁺-driven transporters of the RND family^3,4, enables tissue-patterning activity of the lipid-modified Hedgehog protein by releasing it from tightly -localized sites of embryonic expression^5,6,7,8,9,10. Here we determine a cryo-electron microscopy structure of the mouse protein Dispatched homologue 1 (DISP1), revealing three Na⁺ ions coordinated within a channel that traverses its transmembrane domain. We find that the rate of Hedgehog export is dependent on the Na⁺ gradient across the plasma membrane. The transmembrane channel and Na⁺ binding are disrupted in DISP1-NNN, a variant with asparagine substitutions for three intramembrane aspartate residues that each coordinate and neutralize the charge of one of the three Na⁺ ions. DISP1-NNN and variants that disrupt single Na⁺ sites retain binding to, but are impaired in export of the lipid-modified Hedgehog protein to the SCUBE2 acceptor. Interaction of the amino-terminal signalling domain of the Sonic hedgehog protein (ShhN) with DISP1 occurs via an extensive buried surface area and contacts with an extended furin-cleaved DISP1 arm. Variability analysis reveals that ShhN binding is restricted to one extreme of a continuous series of DISP1 conformations. The bound and unbound DISP1 conformations display distinct Na⁺-site occupancies, which suggests a mechanism by which transmembrane Na⁺ flux may power extraction of the lipid-linked Hedgehog signal from the membrane. Na⁺-coordinating residues in DISP1 are conserved in PTCH1 and other metazoan RND family members, suggesting that Na⁺ flux powers their conformationally driven activities.

^{Dispatched 蛋白与 NPC1 和 PTCH1 胆固醇转运蛋白1,2}以及RND 家族的H ^{+驱动转运蛋白3,4}相关，通过将脂质修饰的 Hedgehog 蛋白从紧密的释放物中释放出来，从而实现脂质修饰 Hedgehog 蛋白的组织模式活性。胚胎表达的局部位点^5,6,7,8,9,10。在这里，我们确定了小鼠蛋白 Dispatched 同源物 1 (DISP1) 的冷冻电子显微镜结构，揭示了在穿过其跨膜域的通道内协调的三个 Na ^+离子。我们发现 Hedgehog 输出速率取决于质膜上的Na ^+梯度。DISP1-NNN 中的跨膜通道和 Na ⁺结合被破坏，DISP1-NNN 是一种用天冬酰胺取代三个膜内天冬氨酸残基的变体，每个残基协调并中和三个 Na ⁺离子之一的电荷。DISP1-NNN 和破坏单个 Na ⁺位点的变体保留与脂质修饰的 Hedgehog 蛋白的结合，但在向 SCUBE2 受体输出脂质修饰的 Hedgehog 蛋白时受到损害。Sonic hidehog 蛋白 (ShhN) 的氨基末端信号结构域与 DISP1 的相互作用通过广泛的埋藏表面区域发生，并与延伸的弗林蛋白酶切割的 DISP1 臂接触。变异性分析表明，ShhN 结合仅限于一系列连续 DISP1 构象的一个极端。结合和未结合的 DISP1 构象显示出不同的 Na ⁺位点占据，这表明跨膜 Na ⁺通量可能为从膜中提取脂质连接的 Hedgehog 信号提供动力。DISP1 中的Na ⁺配位残基在 PTCH1 和其他后生动物 RND 家族成员中是保守的，表明 Na ⁺通量增强了它们的构象驱动活性。