UUKUNIEMI VIRUS MATURATION - IMMUNOFLUORESCENCE MICROSCOPY WITH MONOCLONAL GLYCOPROTEIN-SPECIFIC ANTIBODIES
UUKUNIEMI VIRUS MATURATION - IMMUNOFLUORESCENCE MICROSCOPY WITH MONOCLONAL GLYCOPROTEIN-SPECIFIC ANTIBODIES
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DOI：
10.1128/jvi.51.1.137-146.1984
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发表时间：
1984-01-01
影响因子：
5.4
通讯作者：
PETTERSSON, RF
中科院分区：
文献类型：
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作者：
KUISMANEN, E;BANG, B;PETTERSSON, RF
针对Uukuniemi病毒糖蛋白G1和G2的小鼠单克隆抗体与针对核蛋白（N）的多克隆抗体组合用于研究病毒在感染的鸡胚成纤维细胞和仓鼠肾BHK细胞的高尔基复合体中的成熟。在获得的25个单克隆抗体中，通过免疫印迹和免疫沉淀，10个显示为G1特异性，15个显示为G2特异性。在双染色实验中，与从多克隆兔抗G1-G2抗体获得的染色相比，一些单克隆抗体给出了类似的荧光分布。其他优先染色要么在高尔基复合体或那些在细胞表面的糖蛋白。这可能表明糖蛋白在转运过程中发生了构象变化。Uukuniemi病毒感染导致高尔基复合体膜的空泡化，在那里发生病毒的成熟。 双染色实验与单克隆抗体优先染色的高尔基体相关的病毒糖蛋白和抗N多克隆兔抗血清显示了高尔基体复合体的进行性空泡化和病毒核蛋白在高尔基体区域的积累之间的相关性，这表明高尔基体复合体的形态改变可能是病毒细胞内成熟的先决条件。用衣霉素（一种抑制N-连接糖基化的药物）处理Uukuniemi病毒感染的细胞，导致两种糖蛋白在细胞内位置的积累，显然代表内质网。双染色实验表明，在这些网站的核蛋白的平行积累，表明糖蛋白的局部积累是必需的核蛋白结合到细胞内膜。
Mouse monoclonal antibodies directed against Uukuniemi virus glycoproteins G1 and G2 in combination with polyclonal antibodies against the nucleoprotein (N) were used to study the maturation of the virus in Golgi complexes of infected chicken embryo fibroblasts and hamster kidney BHK cells. Of 25 monoclonal antibodies obtained, 10 were shown to be G1 specific and 15 were shown to be G2 specific by immunoblotting and immunoprecipitation. In double-staining experiments, some of the monoclonal antibodies gave similar distributions of fluorescence as compared with the staining obtained from polyclonal rabbit anti-G1-G2 antibodies. Others preferentially stained either the glycoproteins in the Golgi complex or those at the cell surface. This may indicate that the glycoproteins underwent conformational changes during their transport. Uukuniemi virus infection resulted in the vacuolization of the membranes of Golgi complexes where the maturation of the virus was taking place. Double-staining experiments with monoclonal antibodies which preferentially stained the Golgi-associated viral glycoproteins and with anti-N polyclonal rabbit antiserum showed a correlation between the progressive vacuolization of the Golgi complex and the accumulation of viral nucleoprotein in the Golgi region, suggesting that a morphological alteration of the Golgi complex may be a prerequisite for intracellular maturation of the virus. Treatment of Uukuniemi virus-infected cells with tunicamycin, a drug which inhibits N-linked glycosylation, resulted in the accumulation of both glycoproteins at an intracellular location, apparently representing the endoplasmic reticulum. Double-staining experiments showed a parallel accumulation of nucleoprotein at these sites, indicating that local accumulation of glycoproteins is required for nucleoprotein binding to intracellular membranes.