J Allyn Taylor Centre for Cell Biology, Molecular Brain Research Group, Robarts Research Institute, The University of Western Ontario, 100 Perth Dr, London, ON, N6A 5K8, Canada.
Mol Brain. 2010 Jan 21;3:4. doi: 10.1186/1756-6606-3-4.
Group I metabotropic glutamate receptors (mGluRs) are coupled via Galphaq/11 to the activation of phospholipase Cbeta, which hydrolyzes membrane phospholipids to form inositol 1,4,5 trisphosphate and diacylglycerol. This results in the release of Ca2+ from intracellular stores and the activation of protein kinase C. The activation of Group I mGluRs also results in ERK1/2 phosphorylation. We show here, that the proline-rich tyrosine kinase 2 (Pyk2) interacts with both mGluR1 and mGluR5 and is precipitated with both receptors from rat brain. Pyk2 also interacts with GST-fusion proteins corresponding to the second intracellular loop and the distal carboxyl-terminal tail domains of mGluR1a. Pyk2 colocalizes with mGluR1a at the plasma membrane in human embryonic kidney (HEK293) cells and with endogenous mGluR5 in cortical neurons. Pyk2 overexpression in HEK293 results in attenuated basal and agonist-stimulated inositol phosphate formation in mGluR1 expressing cells and involves a mechanism whereby Pyk2 displaces Galphaq/11 from the receptor. The activation of endogenous mGluR1 in primary mouse cortical neuron stimulates ERK1/2 phosphorylation. Treatments that prevent Pyk2 phosphorylation in cortical neurons, and the overexpression of Pyk2 dominant-negative and catalytically inactive Pyk2 mutants in HEK293 cells, prevent ERK1/2 phosphorylation. The Pyk2 mediated activation of ERK1/2 phosphorylation is also Src-, calmodulin- and protein kinase C-dependent. Our data reveal that Pyk2 couples the activation mGluRs to the mitogen-activated protein kinase pathway even though it attenuates mGluR1-dependent G protein signaling.
I 型代谢型谷氨酸受体（mGluRs）通过 Galphaq/11 与磷脂酶 Cβ的激活偶联，该酶将膜磷脂水解为肌醇 1,4,5 三磷酸和二酰基甘油。这导致细胞内储存的 Ca2+释放和蛋白激酶 C 的激活。I 型 mGluRs 的激活也导致 ERK1/2 的磷酸化。我们在这里表明，富含脯氨酸的酪氨酸激酶 2（Pyk2）与 mGluR1 和 mGluR5 相互作用，并与大鼠脑中的这两种受体沉淀。Pyk2 还与 GST 融合蛋白相互作用，这些融合蛋白对应于 mGluR1a 的第二细胞内环和远端羧基末端尾部结构域。Pyk2 在人胚肾（HEK293）细胞中与 mGluR1a 共定位在质膜上，在皮质神经元中与内源性 mGluR5 共定位。在 HEK293 中过表达 Pyk2 会导致 mGluR1 表达细胞中基础和激动剂刺激的肌醇磷酸盐形成减弱，涉及 Pyk2 将 Galphaq/11 从受体上置换的机制。内源性 mGluR1 在原代小鼠皮质神经元中的激活刺激 ERK1/2 的磷酸化。在皮质神经元中阻止 Pyk2 磷酸化的处理，以及在 HEK293 细胞中过表达 Pyk2 显性负和催化失活的 Pyk2 突变体，可防止 ERK1/2 的磷酸化。Pyk2 介导的 ERK1/2 磷酸化的激活也依赖于Src、钙调蛋白和蛋白激酶 C。我们的数据表明，Pyk2 将 mGluR 的激活与丝裂原活化蛋白激酶途径偶联，尽管它减弱了 mGluR1 依赖性 G 蛋白信号传导。
