Putyrski Mateusz, Schultz Carsten
Cell Biology and Biophysics Unit, European Molecular Biology Laboratory, Meyerhofstr. 1, 69117 Heidelberg, Germany.
Chem Biol. 2011 Sep 23;18(9):1126-33. doi: 10.1016/j.chembiol.2011.07.013.
The selective manipulation of single intracellular-signaling events remains one of the key tasks when studying signaling networks. Here, we demonstrate for the first time the stimulation of FKBP fusions of various subunits of heterotrimeric G proteins by the simple addition of the chemical dimerizer rapamycin. Activation of constitutively active Gα(q), but not its GDP-bound form, leads to sustained oscillations of intracellular calcium and myo-inositol 1,4,5-trisphosphate (InsP(3)) levels in HEK cells, independent of the activation of endogenous Gα(q), in full agreement with the InsP(3)-Ca(2+) cross-coupling model of calcium oscillations. Rapamycin-induced translocation of wild-type Gα(s) to the plasma membrane results in elevated cAMP levels. Activation of rapamycin-inducible Gα(s) or Gβ(1)γ(2) evokes extensive modulation of ATP-induced calcium transients. The results demonstrate that inducible heterotrimeric G protein subunits will provide ways for dissecting G protein-coupled receptor signaling.
在研究信号网络时，对单个细胞内信号事件进行选择性操控仍然是关键任务之一。在此，我们首次证明，通过简单添加化学二聚体雷帕霉素，可刺激异源三聚体G蛋白各亚基的FKBP融合蛋白。组成型活性Gα(q)而非其GDP结合形式的激活，会导致HEK细胞内钙离子和肌醇1,4,5-三磷酸（InsP(3)）水平持续振荡，且与内源性Gα(q)的激活无关，这与钙离子振荡的InsP(3)-Ca(2+)交叉偶联模型完全一致。雷帕霉素诱导野生型Gα(s)转位至质膜会导致cAMP水平升高。雷帕霉素诱导型Gα(s)或Gβ(1)γ(2)的激活会引发ATP诱导的钙瞬变的广泛调节。结果表明，可诱导的异源三聚体G蛋白亚基将为剖析G蛋白偶联受体信号传导提供方法。
