Characterization of the RNA associated with influenza A cytoplasmic inclusions and the interaction of NS1 protein with RNA.
Characterization of the RNA associated with influenza A cytoplasmic inclusions and the interaction of NS1 protein with RNA.
复制标题
与甲型流感细胞质内含物相关的 RNA 的表征以及 NS1 蛋白与 RNA 的相互作用。
DOI：
10.1016/0042-6822(81)90010-6
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发表时间：
1981
期刊：
影响因子：
3.7
通讯作者：
Compans,RW
中科院分区：
文献类型：
--
作者：
Yoshida,T;Shaw,MW;Young,JF;Compans,RW
在感染的BHK 21-F细胞中，发现由甲型流感病毒非结构蛋白（NS 1）形成的次结晶包涵体含有大量的RNA。PAGE分析和寡核苷酸指纹图谱表明，包涵体相关的RNA主要是细胞来源的。通过寡脱氧胸苷酸-纤维素层析和体外翻译对RNA的分析表明，该RNA不是宿主或病毒的信使RNA。纯化后的包涵体用6 Murea和1 MKCl溶解，蔗糖梯度离心分离RNA和NS 1蛋白。当透析去除尿素和KCl时，NS 1蛋白与RNA重新结合。在不存在Mg 2+的情况下得到的复合物似乎比原始NS，夹杂物的尺寸更小，更不均匀，而在10 mMMg 2+的存在下得到的复合物的尺寸与原始NS 1夹杂物相似。无RNA的NS 1蛋白也形成了类似于紧密堆积的NS 1-RNA复合物的紧密堆积结构，表明RNA对于这种相互作用不是必需的。在CsCl中，原始NS 1包涵体、重新结合的NS 1-RNA复合物和无RNA的NS 1蛋白复合物的浮力密度分别为1.347、1.341和1.270 g/ml。结果表明，甲型流感病毒NS 1蛋白具有RNA结合活性，在病毒复制过程中在细胞质中形成含有结合RNA分子的包涵体。
Paracrystalline inclusions formed by influenza A virus nonstructural protein (NS1) in infected BHK21-F cells were found to contain significant amounts of RNA. PAGE analysis and oligonucleotide fingerprint mapping indicated that the inclusion-associated RNA was primarily of cellular origin. Analysis of the RNA by oligodeoxythymidylic acid—cellulose chromatography andin vitrotranslation indicated that the RNA was not host or viral messenger RNA. Purified inclusions were solubilized with 6Murea and 1MKCl, and the NS1protein was separated from the RNA by sucrose gradient centrifugation. When urea and KCl were removed by dialysis, the NS1protein reassociated with the RNA. The resulting complexes obtained in the absence of Mg2+appeared to be smaller and more heterogeneous in size than the original NS, inclusions, whereas complexes obtained in the presence of 10 mMMg2+were similar in size to the original NS1inclusions. RNA-free NS1protein also formed tightly packed structures similar to the tightly packed NS1-RNA complexes, indicating that RNA is not necessary for this interaction. The buoyant densities in CsCl of the original NS1inclusions, the reassociated NS1-RNA complexes, and the RNA-free NS1protein complexes were determined to be 1.347, 1.341, and 1.270 g/ml, respectively. The results indicate that influenza A NS1protein has binding activity for RNA, and forms inclusions containing bound RNA molecules in the cytoplasm during virus replication.