Abstract:
During Hedgehog (Hh) signal transduction in development and disease, the atypical G protein-coupled receptor (GPCR) SMOOTHENED (SMO) communicates with GLI transcription factors by binding the protein kinase A catalytic subunit (PKA-C) and physically blocking its enzymatic activity. Here, we show that GPCR kinase 2 (GRK2) orchestrates this process during endogenous mouse and zebrafish Hh pathway activation in the primary cilium. Upon SMO activation, GRK2 rapidly relocalizes from the ciliary base to the shaft, triggering SMO phosphorylation and PKA-C interaction. Reconstitution studies reveal that GRK2 phosphorylation enables active SMO to bind PKA-C directly. Lastly, the SMO-GRK2-PKA pathway underlies Hh signal transduction in a range of cellular and in vivo models. Thus, GRK2 phosphorylation of ciliary SMO and the ensuing PKA-C binding and inactivation are critical initiating events for the intracellular steps in Hh signaling. More broadly, our study suggests an expanded role for GRKs in enabling direct GPCR interactions with diverse intracellular effectors.
摘要:
在Hedgehog(Hh)信号转导过程中的发育和疾病,非典型G蛋白偶联受体(GPCR)SMOOTHENED(SMO)通过结合蛋白激酶A催化亚基(PKA-C)并物理阻断其酶活性与GLI转录因子通信。这里,我们显示GPCR激酶2(GRK2)在原代纤毛内源性小鼠和斑马鱼Hh通路激活过程中协调这一过程。SMO激活后,GRK2从纤毛基部快速重新定位到轴,触发SMO磷酸化和PKA-C相互作用。重构研究表明,GRK2磷酸化使活性SMO能够直接结合PKA-C。最后,SMO-GRK2-PKA途径是一系列细胞和体内模型中Hh信号转导的基础。因此,纤毛SMO的GRK2磷酸化以及随后的PKA-C结合和失活是Hh信号传导中细胞内步骤的关键启动事件。更广泛地说,我们的研究表明,GRKs在实现与不同细胞内效应物的直接GPCR相互作用方面具有扩大的作用。