Department of Physiology and Neurobiology, University of Connecticut, Storrs, Connecticut 06269, USA.
J Biol Chem. 2011 Jun 24;286(25):22456-68. doi: 10.1074/jbc.M111.236190. Epub 2011 May 3.
Collybistin promotes submembrane clustering of gephyrin and is essential for the postsynaptic localization of gephyrin and γ-aminobutyric acid type A (GABA(A)) receptors at GABAergic synapses in hippocampus and amygdala. Four collybistin isoforms are expressed in brain neurons; CB2 and CB3 differ in the C terminus and occur with and without the Src homology 3 (SH3) domain. We have found that in transfected hippocampal neurons, all collybistin isoforms (CB2(SH3+), CB2(SH3-), CB3(SH3+), and CB3(SH3-)) target to and concentrate at GABAergic postsynapses. Moreover, in non-transfected neurons, collybistin concentrates at GABAergic synapses. Hippocampal neurons co-transfected with CB2(SH3-) and gephyrin developed very large postsynaptic gephyrin and GABA(A) receptor clusters (superclusters). This effect was accompanied by a significant increase in the amplitude of miniature inhibitory postsynaptic currents. Co-transfection with CB2(SH3+) and gephyrin induced the formation of many (supernumerary) non-synaptic clusters. Transfection with gephyrin alone did not affect cluster number or size, but gephyrin potentiated the clustering effect of CB2(SH3-) or CB2(SH3+). Co-transfection with CB2(SH3-) or CB2(SH3+) and gephyrin did not affect the density of presynaptic GABAergic terminals contacting the transfected cells, indicating that collybistin is not synaptogenic. Nevertheless, the synaptic superclusters induced by CB2(SH3-) and gephyrin were accompanied by enlarged presynaptic GABAergic terminals. The enhanced clustering of gephyrin and GABA(A) receptors induced by collybistin isoforms was not accompanied by enhanced clustering of neuroligin 2. Moreover, during the development of GABAergic synapses, the clustering of gephyrin and GABA(A) receptors preceded the clustering of neuroligin 2. We propose a model in which the SH3- isoforms play a major role in the postsynaptic accumulation of GABA(A) receptors and in GABAergic synaptic strength.
胶连蛋白促进胶质纤维酸性蛋白在亚膜区的聚集，对于胶质纤维酸性蛋白和γ-氨基丁酸 A 型（GABA(A)) 受体在海马和杏仁核 GABA 能突触中的突触后定位是必需的。四种胶连蛋白异构体在脑神经元中表达；CB2 和 CB3 在 C 端不同，存在和不存在Src 同源 3（SH3）结构域。我们发现，在转染的海马神经元中，所有胶连蛋白异构体（CB2(SH3+)、CB2(SH3-)、CB3(SH3+)和 CB3(SH3-))都定位于 GABA 能突触后，并在那里聚集。此外，在未转染的神经元中，胶连蛋白也聚集在 GABA 能突触上。与 CB2(SH3-)和胶质纤维酸性蛋白共转染的海马神经元形成了非常大的突触后胶质纤维酸性蛋白和 GABA(A)受体簇（超簇）。这种效应伴随着微小抑制性突触后电流幅度的显著增加。与 CB2(SH3+)和胶质纤维酸性蛋白共转染诱导形成了许多（多余的）非突触簇。单独转染胶质纤维酸性蛋白不会影响簇的数量或大小，但胶质纤维酸性蛋白增强了 CB2(SH3-)或 CB2(SH3+)的聚类效应。与 CB2(SH3-)或 CB2(SH3+)和胶质纤维酸性蛋白共转染不会影响与转染细胞接触的 GABA 能突触前末梢的密度，表明胶连蛋白不是突触形成原。然而，CB2(SH3-)和胶质纤维酸性蛋白诱导的突触超簇伴随着 GABA 能突触前末梢的扩大。胶连蛋白异构体诱导的胶质纤维酸性蛋白和 GABA(A)受体的增强聚类并没有伴随着神经黏附素 2 的增强聚类。此外，在 GABA 能突触的发育过程中，胶质纤维酸性蛋白和 GABA(A)受体的聚类先于神经黏附素 2 的聚类。我们提出了一个模型，其中 SH3-异构体在 GABA(A)受体的突触后聚集和 GABA 能突触强度中起主要作用。
