The morphogen Sonic Hedgehog (SHH) patterns tissues during development by directing cell fates in a concentration-dependent manner. The SHH signal is transmitted across the membrane of target cells by the heptahelical transmembrane protein Smoothened (SMO), which activates the GLI family of transcription factors through a mechanism that is undefined in vertebrates. Using CRISPR-edited null alleles and small-molecule inhibitors, we systematically analyzed the epistatic interactions between SMO and three proteins implicated in SMO signaling: the heterotrimeric G protein subunit Gα_S, the G protein–coupled receptor kinase 2 (GRK2), and the Gα_S-coupled receptor GPR161. Our experiments uncovered a signaling mechanism that modifies the sensitivity of target cells to SHH and consequently changes the shape of the SHH dose-response curve. In both fibroblasts and spinal neural progenitors, the loss of GPR161, previously implicated as an inhibitor of basal SHH signaling, increased the sensitivity of target cells across the entire spectrum of SHH concentrations. Even in cells lacking GPR161, GRK2 was required for SHH signaling, and Gα_s, which promotes the activation of protein Kinase A (PKA), antagonized SHH signaling. We propose that the sensitivity of target cells to Hedgehog morphogens, and the consequent effects on gene expression and differentiation outcomes, can be controlled by signals from G protein–coupled receptors that converge on Gα_s and PKA.

morphogen Sonic Hedgehog (SHH) 在发育过程中通过以浓度依赖性方式指导细胞命运来塑造组织。SHH 信号通过七螺旋跨膜蛋白 Smoothened (SMO) 传递穿过靶细胞的膜，SMO 通过脊椎动物中未定义的机制激活 GLI 转录因子家族。使用 CRISPR 编辑的无效等位基因和小分子抑制剂，我们系统地分析了 SMO 与三种参与 SMO 信号传导的蛋白质之间的上位相互作用：异三聚体 G 蛋白亚基 Gα _S、G 蛋白偶联受体激酶 2 (GRK2) 和GαS _{_}-偶联受体 GPR161。我们的实验揭示了一种信号机制，它改变了靶细胞对 SHH 的敏感性，从而改变了 SHH 剂量反应曲线的形状。在成纤维细胞和脊髓神经祖细胞中，GPR161 的缺失（以前被认为是基础 SHH 信号传导的抑制剂）增加了靶细胞在整个 SHH 浓度范围内的敏感性。即使在缺乏 GPR161 的细胞中，GRK2 也是 SHH 信号传导所必需的，而 Gα _s促进蛋白激酶 A (PKA) 的激活，拮抗 SHH 信号传导。我们提出，靶细胞对 Hedgehog 形态发生素的敏感性，以及由此对基因表达和分化结果的影响，可以由来自 G 蛋白偶联受体的信号控制，这些受体会聚在 Gα 上。_s和 PKA。