Functional analysis of posttranslational cleavage products of the neuron-glia cell adhesion molecule, Ng-CAM.
Functional analysis of posttranslational cleavage products of the neuron-glia cell adhesion molecule, Ng-CAM.
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DOI：
10.1083/jcb.130.3.733
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发表时间：
1995-08
期刊：
影响因子：
--
通讯作者：
Cunningham BA
中科院分区：
文献类型：
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作者：
Burgoon MP;Hazan RB;Phillips GR;Crossin KL;Edelman GM;Cunningham BA
神经元-胶质细胞黏附分子（Ng-CAM）介导神经元间的细胞黏附和神经元与胶质间的细胞黏附；它还促进神经突起的生长。在鸡脑中，Ng- cam被检测为190和210 kD的糖蛋白（Ng- CAM200），翻译后裂解产物为135 kD （F135，包含大部分细胞外区域）和80 kD （F80，包括跨膜和细胞质区域）。为了研究这些成分的功能，我们在小鼠L细胞中表达了Ng-CAM200、F135和F80，并在细菌的pGEX载体中表达了F135和F80作为GST融合蛋白。适当转染的L细胞在其表面表达这两种蛋白；在转染Ng-CAM200和F135的细胞培养基中也发现了F135。除了亲和性结合外，转染了Ng-CAM200和F135的细胞与未转染的L细胞具有异亲和性结合，这表明成纤维细胞上存在Ng-CAM的配体，可能与胶质配体有关。利用转染细胞和融合蛋白的详细研究表明，Ng-CAM的亲同性和异性结合活性都定位在分子的F135片段上。结果还表明，Ng- CAM200的蛋白水解裂解对于其在细胞表面的表达或细胞粘附都不是必需的，并且F135在L细胞上（可能在神经元上）存在一个“锚点”。与细胞结合结果相反，F80而非F135融合蛋白促进了背根神经节细胞的神经突的生长；该活性与F80的FnIII重复序列有关。F135对应的蛋白包含细胞聚集位点，而F80对应的蛋白具有促进神经突生长的能力，这表明蛋白质水解裂解可能是在胚胎发育和神经再生过程中调节Ng-CAM活性的重要事件。
Neuron-glia cell adhesion molecule (Ng-CAM) mediates cell adhesion between neurons homophilically and between neurons and glia heterophilically; it also promotes neurite outgrowth. In the chick brain, Ng-CAM is detected as glycoproteins of 190 and 210 kD (Ng- CAM200) with posttranslational cleavage products of 135 kD (F135, which contains most of the extracellular region) and 80 kD (F80, which includes the transmembrane and the cytoplasmic domains). To examine the functions of each of these components, we have expressed Ng-CAM200, F135, and F80 in murine L cells, and F135 and F80 as GST fusion proteins in the pGEX vector in bacteria. Appropriately transfected L cells expressed each of these proteins on their surfaces; F135 was also found in the media of cells transfected with Ng-CAM200 and F135. In addition to binding homophilically, cells transfected with Ng-CAM200 and F135 bound heterophilically to untransfected L cells, suggesting that there is a ligand for Ng-CAM on fibroblasts that may be related to the glial ligand. Detailed studies using the transfected cells and the fusion proteins indicated that both the homophilic and the heterophilic binding activities of Ng-CAM are localized in the F135 fragment of the molecule. The results also indicated that proteolytic cleavage of Ng- CAM200 is not required either for its expression on the cell surface or for cell adhesion and that there is an "anchor" for F135 on L cells (and presumably on neurons). In contrast to the cell binding results, the F80 but not the F135 fusion protein enhanced the outgrowth of neurites from dorsal root ganglion cells; this activity was associated with the FnIII repeats of F80. The observations that a protein corresponding to F135 contains the cell aggregation sites whereas one corresponding to the F80 has the ability to promote neurite outgrowth suggest that proteolytic cleavage may be an important event in regulating these Ng-CAM activities during embryonic development and neural regeneration.