Li Shuang, Ma Guoqiang, Wang Bing, Jiang Jin
Department of Developmental Biology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA.
Department of Developmental Biology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA. Department of Pharmacology, University of Texas Southwestern Medical Center at Dallas, Dallas, TX 75390, USA.
Sci Signal. 2014 Jul 1;7(332):ra62. doi: 10.1126/scisignal.2005414.
Hedgehog (Hh) is a secreted glycoprotein that binds its receptor Patched to activate the G protein (heterotrimeric guanine nucleotide-binding protein)-coupled receptor-like protein Smoothened (Smo). In Drosophila, protein kinase A (PKA) phosphorylates and activates Smo in cells stimulated with Hh. In unstimulated cells, PKA phosphorylates and inhibits the transcription factor Cubitus interruptus (Ci). We found that in cells exposed to Hh, the catalytic subunit of PKA (PKAc) bound to the juxtamembrane region of the carboxyl terminus of Smo. PKA-mediated phosphorylation of Smo further enhanced its association with PKAc to form stable kinase-substrate complexes that promoted the PKA-mediated transphosphorylation of Smo dimers. We identified multiple basic residues in the carboxyl terminus of Smo that were required for interaction with PKAc, Smo phosphorylation, and Hh pathway activation. Hh induced a switch from the association of PKAc with a cytosolic complex of Ci and the kinesin-like protein Costal2 (Cos2) to a membrane-bound Smo-Cos2 complex. Thus, our study uncovers a previously uncharacterized mechanism for regulation of PKA activity and demonstrates that the signal-regulated formation of kinase-substrate complexes plays a central role in Hh signal transduction.
刺猬蛋白（Hh）是一种分泌型糖蛋白，它与受体patched结合以激活G蛋白（异源三聚体鸟嘌呤核苷酸结合蛋白）偶联受体样蛋白 smoothened（Smo）。在果蝇中，蛋白激酶A（PKA）在受Hh刺激的细胞中磷酸化并激活Smo。在未受刺激的细胞中，PKA磷酸化并抑制转录因子间断翅脉（Ci）。我们发现，在暴露于Hh的细胞中，PKA的催化亚基（PKAc）与Smo羧基末端的近膜区域结合。PKA介导的Smo磷酸化进一步增强了其与PKAc的结合，形成稳定的激酶-底物复合物，促进了PKA介导的Smo二聚体的转磷酸化。我们在Smo的羧基末端鉴定出多个碱性残基，这些残基是与PKAc相互作用、Smo磷酸化和Hh信号通路激活所必需的。Hh诱导了PKAc从与Ci和类驱动蛋白Costal2（Cos2）的胞质复合物的结合转变为与膜结合的Smo-Cos2复合物的结合。因此，我们的研究揭示了一种以前未被描述的PKA活性调节机制，并证明激酶-底物复合物的信号调节形成在Hh信号转导中起核心作用。
