Connexin47, connexin29 and connexin32 co-expression in oligodendrocytes and Cx47 association with zonula occludens-1 (Zo-1) in mouse brain
Connexin47, connexin29 and connexin32 co-expression in oligodendrocytes and Cx47 association with zonula occludens-1 (Zo-1) in mouse brain
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DOI：
10.1016/j.neuroscience.2004.03.063
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发表时间：
2004-01-01
期刊：
影响因子：
3.3
通讯作者：
Nagy, JI
中科院分区：
文献类型：
--
作者：
Li, X;Ionescu, AV;Nagy, JI
已知哺乳动物CNS中神经胶质细胞之间的间隙连接含有几种连接蛋白（Cx），包括星形胶质细胞-星形胶质细胞连接处的Cx 26、Cx 30和Cx 43，以及星形胶质细胞-少突胶质细胞连接处的少突胶质细胞侧的Cx 29和Cx 32。最近的报道表明，少突胶质细胞也表达Cx47，促使目前的研究Cx47的定位和其他胶质细胞连接蛋白在小鼠中枢神经系统的关系。鉴于越来越多的连接蛋白报道直接与支架蛋白zonula occludens-1（ZO-1）相互作用，我们研究了ZO-1的表达和Cx47/ZO-1在脑，脊髓和Cx47转染HeLa细胞中的相互作用能力。从不同脑区免疫荧光标记的超过9000个少突胶质细胞的计数中，发现几乎所有这些细胞都表达Cx 29、Cx 32和Cx47。少突胶质细胞胞体显示强大的Cx47免疫阳性斑点，共定位与Cx 32和Cx43的点状标记。冷冻断裂复制免疫金标记，Cx47是丰富的少突胶质细胞/星形胶质细胞间隙连接的少突胶质细胞侧。通过免疫荧光，标记为Cx47沿着髓鞘纤维稀疏，在大多数脑区，而Cx 29和Cx 32被发现是集中沿着这些纤维。通过免疫金标记，Cx47被发现在许多连接髓鞘和星形胶质细胞的小缝隙连接中，但不在髓鞘的更深层中。脑亚细胞分级显示缺乏Cx47富集的髓鞘组分，但含有Cx 32和Cx 29的富集。少突胶质细胞ZO-1免疫阳性，并显示几乎所有的Cx47/ZO-1共定位。发现ZO-1与Cx47共免疫沉淀，并且下拉测定表明Cx47与ZO-1的第二PDZ结构域结合。我们的研究结果表明，Cx47的广泛表达的少突胶质细胞，但与分布模式的相对水平相反的丰富的Cx 29的髓鞘和Cx 29的少突胶质细胞胞体。此外，我们的研究结果表明ZO-1在星形胶质细胞与少突胶质细胞间隙连接的少突胶质细胞侧具有支架和/或调节作用。(C)2004年IBRO。由爱思唯尔有限公司出版。保留所有权利。
Gap junctions between glial cells in mammalian CNS are known to contain several connexins (Cx), including Cx26, Cx30 and Cx43 at astrocyte-to-astrocyte junctions, and Cx29 and Cx32 on the oligodendrocyte side of astrocyte-to-oligodendrocyte junctions. Recent reports indicating that oligodendrocytes also express Cx47 prompted the present studies of Cx47 localization and relationships to other glial connexins in mouse CNS. In view of the increasing number of connexins reported to interact directly with the scaffolding protein zonula occludens-1 (ZO-1), we investigated ZO-1 expression and Cx47/ZO-1 interaction capabilities in brain, spinal cord and Cx47-transfected HeLa cells. From counts of over 9000 oligodendrocytes labeled by immunofluorescence in various brain regions, virtually all of these cells were found to express Cx29, Cx32 and Cx47. Oligodendrocyte somata displayed robust Cx47-immunopositive puncta that were colocalized with punctate labeling for Cx32 and Cx43. By freeze-fracture replica immunogold labeling, Cx47 was abundant on the oligodendrocyte-side of oligodendrocyte/astrocyte gap junctions. By immunofluorescence, labeling for Cx47 along myelinated fibers was sparse in most brain regions, whereas Cx29 and Cx32 were previously found to be concentrated along these fibers. By immunogold labeling, Cx47 was found in numerous small gap junctions linking myelin to astrocytes, but not within deeper layers of myelin. Brain subcellular fractionation revealed a lack of Cx47 enrichment in myelin fractions, which nevertheless contained an enrichment of Cx32 and Cx29. Oligodendrocytes were immunopositive for ZO-1, and displayed almost total Cx47/ZO-1 co-localization. ZO-1 was found to co-immunoprecipitate with Cx47, and pull-down assays indicated binding of Cx47 to the second PDZ domain of ZO-1. Our results indicate widespread expression of Cx47 by oligodendrocytes, but with a distribution pattern in relative levels inverse to the abundance of Cx29 in myelin and paucity of Cx29 in oligodendrocyte somata. Further, our findings suggest a scaffolding and/or regulatory role of ZO-1 at the oligodendrocyte side of astrocyte-to-oligodendrocyte gap junctions. (C) 2004 IBRO. Published by Elsevier Ltd. All rights reserved.