GENOMIC RNAS OF INFLUENZA-VIRUSES ARE HELD IN A CIRCULAR CONFORMATION IN VIRIONS AND IN INFECTED-CELLS BY A TERMINAL PANHANDLE
GENOMIC RNAS OF INFLUENZA-VIRUSES ARE HELD IN A CIRCULAR CONFORMATION IN VIRIONS AND IN INFECTED-CELLS BY A TERMINAL PANHANDLE
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DOI：
10.1073/pnas.84.22.8140
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发表时间：
1987-11-01
影响因子：
11.1
通讯作者：
PALESE, P
中科院分区：
文献类型：
--
作者：
HSU, MT;PARVIN, JD;PALESE, P
应用补骨脂素交联方法发现流感病毒粒子中的病毒RNA片段呈环状构象。用补骨脂素试剂4' -氨基甲基三氧沙兰（AMT）处理病毒后，纯化RNA的电子显微镜显示出长度与单个片段对应的圆圈。使用聚丙烯酰胺凝胶的RNA印迹分析表明，amt处理的病毒的RNA迁移速度较慢，与单链环一致。此外，核酸酶S1保护实验表明，RNA片段的末端形成一个大约15个碱基对长的狭长。这种结构与所有流感病毒rna末端所观察到的部分序列互补性一致。通过RNA印迹分析，在流感病毒感染细胞的早期和晚期也发现了病毒感RNA的环状结构。当主要的转录产物是信息RNA时，环状RNA是优势种。这一发现和mRNA合成的终止信号直接靠近panhandle的观察表明，模板病毒RNA中的panhandle是促进mRNA合成的顺式调控信号，而不是正义模板。此外，由于长柄在病毒粒子中高浓度存在，我们认为它是包装所必需的，并且感染后的输入RNA具有合成初级转录本的适当构象。
The viral RNA segments in influenza virions were shown to be circular in conformation by using psoralen crosslinking methods. Electron microscopy of purified RNA following treatment of virus with the psoralen reagent 4''-aminomethyltrioxsalen (AMT) revealed circles with lengths corresponding to the individual segments. RNA blot analysis using polyacrylamide gels demonstrated that RNA from AMT-treated virus had a slowed migration, consistent with it being a single-stranded circle. Furthermore, nuclease S1 protection assays indicated that the termini of the RNA segments form an approximately 15-base-pair-long panhandle. This structure is consistent with the partial sequence complementarity that has been observed for the termini of all influenza virus RNAs. By RNA blot analysis, circular structures of viral sense RNA were also found in influenza virus-infected cells at early and late time points. The circular RNA was the predominant species at the time when the major transcription product is message RNA. This finding and the observation that the termination signal for mRNA synthesis directly abuts the panhandle suggest that a panhandle in the template viral RNA is a cis regulatory signal promoting the synthesis of mRNA instead of plus-sense template. Also, since the panhandle is present in high concentration in virions, we suggest that it is required for packaging and that the input RNA after infection is in the proper conformation for synthesis of primary transcripts.