NMDA receptor GluRε/NR2 Subunits are essential for postsynaptic localization and protein stability of GluRζ1/NR1 subunit
NMDA receptor GluRε/NR2 Subunits are essential for postsynaptic localization and protein stability of GluRζ1/NR1 subunit
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DOI：
10.1523/jneurosci.1261-04.2004
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发表时间：
2004-08-18
影响因子：
5.3
通讯作者：
Sakimura, K
中科院分区：
文献类型：
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作者：
Abe, M;Fukaya, M;Sakimura, K
在NMDA受体中，GluRzeta 1/NR 2亚基严格需要GluRzeta 1/NR 1亚基从内质网（ER）离开到体外细胞表面和体内突触后，而GluRzeta 1亚基在体外已被证明是C末端依赖性的自表面递送。为了测试这是否会导致体内神经元中C末端依赖性的自我突触后表达，我们研究了缺乏两个主要GluRzeta亚基GluRzeta 1亚基的小脑颗粒细胞，GluRzeta 1/NR 2A和GluRzeta 3/NR 2C。在突变的小脑中，含有C2（GluRzeta 1-C2）或C2'（GluRzeta 1-C2'）盒的GluRzeta 1亚基的突触标记在苔藓纤维-颗粒细胞突触处减少到突触外水平。这种损失并不伴随着转录和翻译水平的降低，突触外标记的增加或ER的积累。定量免疫印迹显示GluRzeta 1-C2和GluRzeta 1-C2 '的突变小脑显著减少。在突触后密度（PSD）部分中观察到最严重的缺陷：相对于野生型水平，GluRzeta 1-C2的突变水平为12.3 +/- 3.3%，GluRzeta 1-C2 '为17.0 +/- 4.6%。携带C1盒的GluRzeta 1亚基（GluRzeta 1-C1）虽然小脑含量低，但在突变PSD组分中也降低至12.7 +/- 3.5%。考虑到突变小脑中微量的其他GluRzeta亚基，因此严重的减少代表GluRzeta 1亚基本身实际上无法在突触后位点积累，无论C-末端形式如何。通过蛋白质周转率分析，GluRzeta 1亚基的降解在突变小脑中加速，对于携带C2盒的小脑尤其迅速。因此，GluRzeta 1亚基的伴随表达对于GluRzeta 1亚基的突触后定位和蛋白质稳定性至关重要。
In NMDA receptors, GluRepsilon/NR2 subunits strictly require the GluRzeta1/NR1 subunit to exit from endoplasmic reticulum ( ER) to the cell surface in vitro and to the postsynapse in vivo, whereas Cterminus-dependent self-surface delivery has been demonstrated for the GluRzeta1 subunit in vitro. To test whether this leads to C terminus-dependent self-postsynaptic expression in neurons in vivo, we investigated the GluRzeta1 subunit in cerebellar granule cells lacking two major GluRepsilon subunits, GluRepsilon1/NR2Aand GluRepsilon3/NR2C. In the mutant cerebellum, synaptic labeling for the GluRzeta1 subunit containing the C2 (GluRzeta1-C2) or C2' (GluRzeta1-C2') cassette was reduced at mossy fiber-granule cell synapses to the extrasynaptic level. The loss was not accompanied by decreased transcription and translation levels, increased extrasynaptic labeling, or ER accumulation. Quantitative immunoblot revealed substantial reductions in the mutant cerebellum of GluRzeta1-C2 and GluRzeta1-C2'. The most severe deficit was observed in the postsynaptic density (PSD) fraction: mutant levels relative to the wild-type level were 12.3 +/- 3.3% for GluRzeta1-C2 and 17.0 +/- 4.6% for GluRzeta1-C2'. The GluRzeta1 subunit carrying the C1 cassette (GluRzeta1-C1) was, although low in cerebellar content, also reduced to 12.7 +/- 3.5% in the mutant PSD fraction. Considering a trace amount of other GluRepsilon subunits in the mutant cerebellum, the severe reductions thus represent that the GluRzeta1 subunit, by itself, is virtually unable to accumulate at postsynaptic sites, regardless of C-terminal forms. By protein turnover analysis, the degradation of the GluRzeta1 subunit was accelerated in the mutant cerebellum, being particularly rapid for that carrying the C2 cassette. Therefore, accompanying expression of GluRepsilon subunits is essential for postsynaptic localization and protein stability of the GluRzeta1 subunit.