Rapid and quantitative evaluation of vaccinia virus-induced host shutoff using newly generated cell lines stably expressing secreted Gaussia luciferase.
Rapid and quantitative evaluation of vaccinia virus-induced host shutoff using newly generated cell lines stably expressing secreted Gaussia luciferase.
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DOI：
10.1002/jmv.27773
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发表时间：
2022-08
影响因子：
12.7
通讯作者：
Yang, Zhilong
中科院分区：
文献类型：
--
作者：
Molina, Joshua A.;Yang, Zhilong
宿主关闭，其特征在于细胞蛋白质合成的整体下降，通常在许多病毒感染中观察到，包括牛痘病毒。测量宿主关闭的经典方法包括使用放射性或非放射性探针标记新合成的蛋白质，然后进行放射自显影或十二烷基硫酸钠聚丙烯酰胺凝胶电泳（SDS-PAGE）以解析蛋白质用于后续检测。虽然这些都是高度可靠的方法，但它们耗时耗力。在这里，我们产生了两种稳定表达分泌型Gaussia荧光素酶的细胞系。这些报告细胞允许从单个感染样品快速、定量和连续监测宿主关闭。我们使用报告细胞系评估了野生型和各种突变型牛痘病毒诱导的宿主关闭。结果验证了报告细胞的效用，并定量表征了牛痘病毒在复制的不同阶段诱导的宿主关闭。值得注意的是，结果还表明了其他主要的未识别的VACV关闭因素。本研究为研究宿主关闭提供了新的工具。报告细胞也适用于临床分离病毒的高通量设置和快速检测。结合经典的方法，这一工具将大大有助于理解病毒诱导的主机关闭，和蛋白质合成关闭所造成的其他生理相关的压力。
Host shutoff, characterized by a global decline of cellular protein synthesis, is commonly observed in many viral infections, including vaccinia virus. Classic methods measuring host shutoff include the use of radioactive or non-radioactive probes to label newly synthesized proteins followed by radioautography or sodium dodecyl-sulfate polyacrylamide gel electrophoresis (SDS-PAGE) to resolve the proteins for follow-up detection. While these are highly reliable methods, they are time- and labor-consuming. Here we generated two cell lines stably expressing secreted Gaussia luciferase. These reporter cells allow rapid, quantitative, and consecutive monitoring of host shutoff from a single infection sample. We evaluated host shutoff induced by wild-type and various mutant vaccinia viruses using the reporter cell lines. The results validated the utilities of the reporter cells and quantitatively characterized vaccinia virus-induced host shutoff at different stages of replication. Notably, the results also indicated additional major unidentified VACV shutoff factors. Our study provides new tool to study host shutoff. The reporter cells are also suitable for high throughput settings and rapid testing of clinically isolated viruses. In combination with classical methods, this tool will greatly facilitate understanding of virus-induced host shutoff, and protein synthesis shutoff caused by other physiologically relevant stresses.
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DOI：
10.3390/v6072735
发表时间：
2014-07-17
期刊：
Viruses
影响因子：
--
作者：
Altenburg AF;Kreijtz JH;de Vries RD;Song F;Fux R;Rimmelzwaan GF;Sutter G;Volz A
通讯作者：
Volz A
影响因子：
5.4
作者：
Liu, Ruikang;Moss, Bernard
通讯作者：
Moss, Bernard
影响因子：
7.4
作者：
Maguire, Casey A.;Deliolanis, Nikolaos C.;Pike, Lisa;Niers, Johanna M.;Tjon-Kon-Fat, Lee-Ann;Sena-Esteves, Miguel;Tannous, Bakhos A.
通讯作者：
Tannous, Bakhos A.
影响因子：
6.4
作者：
通讯作者：
--
影响因子：
5.4
作者：
Meade, Nathan;King, Melvin;Walsh, Derek
通讯作者：
Walsh, Derek