Neuropharmacological studies of clustering molecules expressing in the excitatory synapses

已经克隆并表征了椎间盘大（dlg）肿瘤抑制蛋白的人同源物NE-dlg（神经元和内分泌-dlg）（K. Makino等人，Oncogene，14，2425，1997）。NE-dlg，也称为SAP 102，与NMDA受体2B（NR 2B）相互作用以使受体聚集在突触后密度中。我们首先证明了NE-dlg与钙调蛋白和PSD-95在体外相互作用（J. Biol. Chem.，274，5782，1999）。我们还在大脑中发现了一种相互作用的蛋白质，p51-Nedasin。P51-Nedasin干扰NE-dlg和NR 2B之间的结合（J.Biol.Chem.274，32204，1999）。接下来，我们证明了在体外和原位磷酸化NE-dlg的钙调素激酶II在突触后密度组分和培养的大鼠海马神经元。在海马长时程增强后也观察到NE-dlg磷酸化增加。用CaM激酶抑制剂KN 93处理可阻止磷酸化增加，但蛋白激酶C抑制剂不能。我们评估了NE-dlg磷酸化的功能相关性。CaM激酶II依赖的NE-dlg磷酸化阻止其与NR 2B结合，但促进NR 2B和NR 1在突触后致密物中的积累。关于NE-dlg的胞质定位，CaM激酶II对NE-dlg的磷酸化可能参与NR 2B和/或NR 1向突触后致密物的转运。我们最近发现，CaM激酶II也参与多巴胺D2受体从高尔基体转运到神经元的质膜，其中支架蛋白如NE-dlg可能是其转运机制的基础。

A human homologue to the discs large (dlg) tumor suppresser protein, NE-dlg (neuronal and endocrine-dlg) has been cloned and characterized (K. Makino et al., Oncogene, 14, 2425, 1997). NE-dlg, also named SAP102, interacts with the NMDA receptor 2B (NR2B) to cluster the receptors in the postsynaptic density. We first documented that NE-dlg interacts with calmodulin and PSD-95 in vitro (J. Biol. Chem., 274, 5782, 1999). We also found an interacting protein, p51-Nedasin, in the brain. The P51-Nedasin interferes the association between NE-dlg and NR2B (J. Biol. Chem. 274, 32204, 1999). We next demonstrated in vitro and in situ phosphorylation of NE-dlg by CaM kinase II in the postsynaptic density fractions and in cultured rat hippocampal neurons. An increased phosphorylation of NE-dlg was also observed following hippocampal long-term potentiation. The increased phosphorylation was prevented by treatment with KN93, a CaM kinase inhibitor but not by protein kinase C inhibitors. We assessed the functional relevance of NE-dlg phosphorylation. CaM kinase Il-dependent NE-dlg phosphorylation prevented its binding to NR2B but promoted the accumulation of NR2B and NR1 in the postsynaptic density. As concerning the cytoplasmic localization of NE-dlg, the phosphorylation of NE-dlg by CaM kinase II is possibly involved in the transport of NR2B and/or NR1 into the postsynaptic density. We recently found that CaM kinase II is also involved in transport of dopamine D2 receptors from Golgi apparatus to the plasma membrane in the neurons, in which a scaffold protein like NE-dlg may underlie its transport mechanisms.