SPECIFICITY OF ROUS-SARCOMA VIRUS NUCLEOCAPSID PROTEIN IN GENOMIC RNA PACKAGING
SPECIFICITY OF ROUS-SARCOMA VIRUS NUCLEOCAPSID PROTEIN IN GENOMIC RNA PACKAGING
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DOI：
10.1128/jvi.66.8.4662-4670.1992
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发表时间：
1992-08-01
影响因子：
5.4
通讯作者：
SPAHR, PF
中科院分区：
文献类型：
--
作者：
DUPRAZ, P;SPAHR, PF
定点诱变已经表明劳斯肉瘤病毒（RSV）的核衣壳（NC）蛋白是病毒RNA的包装和二聚化所需的。然而，还不能在体内或体外证明NC与病毒RNA序列的特异性结合。为了确定病毒RNA的特异性包装是否由NC在体内介导，我们构建了携带莫洛尼鼠白血病病毒（MoMuLV）序列的RSV突变体。将MoMuLV的单独NC编码区、psi-RNA包装序列或NC和psi序列两者分别取代全长RSV克隆的相应区域，以产生嵌合质粒pAPrcMNC、pAPrc-psi-M或pAPrcM-psi-M。此外，测试了其中NC完全缺失的RSV突变体作为对照。转染后，每种嵌合突变体产生含有加工过的核心蛋白的病毒颗粒，但无感染性。因此，MoMuLV NC可以在成熟病毒体的组装和释放中功能性地替代RSV NC，但在感染性方面不能。令人惊讶的是，全缺失突变体在病毒释放中显示出强烈的阻断作用，表明NC参与病毒组装。突变体PrcMNC包装的RSV RNA比野生型少50- 100倍;在共转染实验中，MoMuLV RNA优先包装。这一结果表明，在病毒组装过程中，病毒RNA的特异性识别至少部分涉及NC蛋白。
Site-directed mutagenesis has shown that the nucleocapsid (NC) protein of Rous sarcoma virus (RSV) is required for packaging and dimerization of viral RNA. However, it has not been possible to demonstrate, in vivo or in vitro, specific binding of viral RNA sequences by NC. To determine whether specific packaging of viral RNA is mediated by NC in vivo, we have constructed RSV mutants carrying sequences of Moloney murine leukemia virus (MoMuLV). Either the NC coding region alone, the psi-RNA packaging sequence, or both the NC and psi-sequences of MoMuLV were substituted for the corresponding regions of a full-length RSV clone to yield chimeric plasmid pAPrcMNC, pAPrc-psi-M, or pAPrcM-psi-M, respectively. In addition, a mutant of RSV in which the NC is completely deleted was tested as a control. Upon transfection, each of the chimeric mutants produced viral particles containing processed core proteins but were noninfectious. Thus, MoMuLV NC can replace RSV NC functionally in the assembly and release of mature virions but not in infectivity. Surprisingly, the full-deletion mutant showed a strong block in virus release, suggesting that NC is involved in virus assembly. Mutant PrcMNC packaged 50- to 100-fold less RSV RNA than did the wild type; in cotranfection experiments, MoMuLV RNA was preferentially packaged. This result suggests that the specific recognition of viral RNA during virus assembly involves, at least in part, the NC protein.