Protein Kinase C Promotes N-Methyl-d-aspartate (NMDA) Receptor Trafficking by Indirectly Triggering Calcium/Calmodulin-dependent Protein Kinase II (CaMKII) Autophosphorylation

Jing-Zhi Yan,Zhuo Xu,Si-Qiang Ren,Bin Hu,Wen Yao,Shan-Hui Wang,Su-Yi Liu,Wei Lu

doi:10.1074/jbc.m110.192708

Abstract

Regulation of neuronal NMDA receptor (NMDAR) is critical in synaptic transmission and plasticity. Protein kinase C (PKC) promotes NMDAR trafficking to the cell surface via interaction with NMDAR-associated proteins (NAPs). Little is known, however, about the NAPs that are critical to PKC-induced NMDAR trafficking. Here, we showed that calcium/calmodulin-dependent protein kinase II (CaMKII) could be a NAP that mediates the potentiation of NMDAR trafficking by PKC. PKC activation promoted the level of autophosphorylated CaMKII and increased association with NMDARs, accompanied by functional NMDAR insertion, at postsynaptic sites. This potentiation, along with PKC-induced long term potentiation of the AMPA receptor-mediated response, was abolished by CaMKII antagonist or by disturbing the interaction between CaMKII and NR2A or NR2B. Further mutual occlusion experiments demonstrated that PKC and CaMKII share a common signaling pathway in the potentiation of NMDAR trafficking and long-term potentiation (LTP) induction. Our results revealed that PKC promotes NMDA receptor trafficking and induces synaptic plasticity through indirectly triggering CaMKII autophosphorylation and subsequent increased association with NMDARs.

Highlights

Increases the NMDA channel opening rate and delivers new NMDA receptor (NMDAR) to the plasma membrane through regulated exocytosis [3, 4]
Using a Western blot assay of the Triton X-100-insoluble fraction of hippocampal tissue [12], which roughly represents the subcellular fraction at postsynaptic sites [12, 29], we found a dramatic elevation of postsynaptic expression of NR2A, NR2B, autophosphorylated calmodulin-dependent protein kinase II (CaMKII), and CaMKII upon Protein kinase C (PKC) activation by 15-min phorbol 12-myristate 13-acetate (PMA) treatment (NR2A, 1.79 Ϯ 0.17, n ϭ 5, p Ͻ 0.01; NR2B, 1.60 Ϯ 0.07, n ϭ 5, p Ͻ 0.01; p-CaMKII, 1.82 Ϯ 0.16, n ϭ 5, p Ͻ 0.01; CaMKII, 1.76 Ϯ 0.11, n ϭ 5, p Ͻ 0.01; ANOVA least significant difference (LSD) test; Fig. 1, A and B)
Consistent with the above findings using Western blot and co-IP, we found that Tat-NR2A completely blocked the increase of postsynaptic NR2A and CaMKII (NR2A, 1.00 Ϯ 0.05, n ϭ 12, p Ͼ 0.05; CaMKII, 1.00 Ϯ 0.04, n ϭ 15, p Ͼ 0.05) as well as enhanced colocalization between NR2A and CaMKII at postsynaptic sites upon PKC activation by PMA (0.92 Ϯ 0.03, n ϭ 12, p Ͼ 0.05 compared with control, one-way ANOVA LSD test; Fig. 6, A and C) but partially blocked NR2B and the colocalization of NR2B and CaMKII (NR2B, 1.16 Ϯ 0.03, n ϭ 10, p Ͻ 0.05; NR2B/ CaMKII, 1.14 Ϯ 0.03, n ϭ 10, p Ͻ 0.05 compared with control)