N-LINKED GLYCOPROTEIN-SYNTHESIS AND TRANSPORT DURING G1 ARE NECESSARY FOR ASTROCYTIC PROLIFERATION
N-LINKED GLYCOPROTEIN-SYNTHESIS AND TRANSPORT DURING G1 ARE NECESSARY FOR ASTROCYTIC PROLIFERATION
复制标题
DOI：
10.1002/jnr.490260404
复制
发表时间：
1990-08-01
影响因子：
4.2
通讯作者：
VOLPE, JJ
中科院分区：
文献类型：
--
作者：
ISHII, S;VOLPE, JJ
采用选择性分离技术从新生大鼠大脑神经胶质原代混合培养物中分离纯化星形胶质细胞，研究其对星形胶质细胞增殖的影响。通过从培养上清液中去除胎牛血清(FCS)，首先诱导细胞处于静止状态，2天后细胞同步；通过将血清返回培养液，逆转静止状态，12-24小时后DNA合成显著增加。2-脱氧葡萄糖是二醇连接的寡糖和N-连接的糖蛋白生物合成的抑制剂，它不仅阻止了细胞周期中G1期糖蛋白生物合成的增加，而且还阻止了随后在S期的DNA合成的爆发。在培养基中加入甘露糖可阻止脱氧葡萄糖对糖蛋白和DNA合成的抑制。这些数据表明，在星形胶质细胞中，DOLICHOL连接的糖蛋白合成和DNA合成之间存在着必然的关系。为了确定新合成的糖蛋白转运到质膜以并入或分泌是否对DNA合成和星形胶质细胞增殖是必需的，我们研究了莫能菌素处理的细胞，莫能菌素是一种单价阳离子载体，也是糖蛋白细胞内运输的抑制剂。同步化星形胶质细胞在血清回流后12小时内加入莫能菌素，可阻止细胞进入S期和细胞增殖；在S期开始后的前12小时加入莫能菌素，对通过S期的进展没有影响。凝集素染色结合荧光显微镜观察表明，莫能菌素处理的细胞不能将细胞内的糖蛋白转运到质膜。这些数据提示，星形胶质细胞在G1期和S期细胞周期中所需的糖蛋白必须转运到细胞表面。这些蛋白质的性质，例如，是质膜成分还是分泌性糖蛋白，以及它们的特定功能仍有待阐明。
The proliferation of astrocytes, purified by a selective detachment technique from mixed glial primary cultures derived from newborn rat cerebrum, was studied. The cells were synchronized by first inducing a quiescent state by removing fetal calf serum (FCS) from the culture medium for 2 days; reversal of the quiescent state by return of serum to the culture medium caused a marked increase in DNA synthesis 12-24 hr later. 2-Deoxyglucose, an inhibitor of dolichol-linked oligosaccharide and thereby N-linked glycoprotein biosynthesis, prevented not only an increase in glycoprotein biosynthesis in G1 phase of the cell cycle but also the burst of DNA synthesis that followed during S phase. Addition of mannose to the culture medium prevented the inhibitions by deoxyglucose of both glycoprotein and DNA syntheses. These data indicated an obligatory relationship in astrocytes between dolichol-linked glycoprotein synthesis and DNA synthesis. To determine whether transport of the newly synthesized glycoproteins to the plasma membrane for incorporation therein or for secretion were necessary for DNA synthesis and astrocytic proliferation, we studied cells treated with monensin, an ionophore for monovalent cations, and an inhibitor of intracellular transport of glycoproteins. The presence of monensin in the first 12 hr after repletion of serum to synchronized astrocytes prevented progression to the S phase and cell proliferation; addition of monensin after the first 12 hr, at the onset of the S phase, had no effect on progression through S phase. Lectin-staining methods combined with fluorescence microscopy demonstrated in monensin-treated cells failure to intracellular glycoproteins to be transported to the plasma membrane. These data suggest that the glycoproteins synthesized during G1 and required for traverse of astrocytes through G1 and the S phase of the cell cycle must be transported to the cell surface. The nature of these proteins, e.g., whether plasma membrane components or secretory glycoproteins, and their specific functions remain to be elucidated.