Four proteins processed from the replicase gene polyprotein of mouse hepatitis virus colocalize in the cell periphery and adjacent to sites of virion assembly
Four proteins processed from the replicase gene polyprotein of mouse hepatitis virus colocalize in the cell periphery and adjacent to sites of virion assembly
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DOI：
10.1128/jvi.74.7.3379-3387.2000
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发表时间：
2000-04-01
影响因子：
5.4
通讯作者：
Denison, MR
中科院分区：
文献类型：
--
作者：
Bost, AG;Carnahan, RH;Denison, MR
冠状病毒小鼠肝炎病毒（MHV）的复制酶基因（基因1）编码两种共氨基末端多聚蛋白，据推测这些多聚蛋白包含病毒RNA合成所需的所有病毒编码蛋白。这些多聚蛋白被病毒蛋白酶共翻译加工成至少15种成熟蛋白，包括4种预测分子量小于25 kDa的裂解产物，它们加起来构成从开放阅读框1a翻译的最终59 kDa的蛋白质。针对4个不同结构域的单特异性抗体在MHV - A59感染的DBT细胞中检测到10、12和15 kDa的蛋白质（p1a - 10、p1a - 12和p1a - 15），此外还有先前已鉴定的22 kDa蛋白质（p1a - 22）。当通过免疫荧光激光共聚焦显微镜检测感染细胞时，在离散的病灶中检测到p1a - 10、 - 22、 - 12和 - 15，这些病灶在核周区域明显，但也广泛分布在整个细胞质中。双标记实验表明，大多数p1a - 22在复制复合物中与解旋酶、核衣壳和3C样蛋白酶以及p1a - 10、 - 12和 - 15共定位。p1a - 22也在复制复合物附近的单独病灶中被检测到。大多数含有基因1蛋白的复合物与M装配蛋白的积累位点不同。然而，在核周区域，基因1蛋白和核衣壳与M蛋白定位位点相互交错。这些结果表明，已知参与RNA合成的复合物包含多种基因1蛋白，并在假定的病毒体装配位点与结构蛋白密切相关。
The replicase gene (gene 1) of the coronavirus mouse hepatitis virus (MHV) encodes two co-amino-terminal polyproteins presumed to incorporate all the virus-encoded proteins necessary for viral RNA synthesis. The polyproteins are cotranslationally processed by viral proteinases into at least 15 mature proteins, including four predicted cleavage products of less than 25 kDa that together would comprise the final 59 kDa of protein translated from open reading frame la. Monospecific antibodies directed against the four distinct domains detected proteins of 10, 12, and 15 kDa (p1a-10, p1a-12, and p1a-15) in MHV-A59-infected DBT cells, in addition to a previously identified 22-kDa protein (p1a-22). When infected cells were probed by immunofluorescence laser confocal microscopy, p1a-10, -22, -12, and -15 were detected in discrete foci that were prominent in the perinuclear region but were widely distributed throughout the cytoplasm as well. Dual-labeling experiments demonstrated colocalization of the majority of p1a-22 in replication complexes with the helicase, nucleocapsid, and 3C-like proteinase, as well as with p1a-10, -12, and -15. p1a-22 was also detected in separate foci adjacent to the replication complexes. The majority of complexes containing the gene 1 proteins were distinct from sites of accumulation of the M assembly protein. However, in perinuclear regions the gene 1 proteins and nucleocapsid were intercalated with sites of M protein localization. These results demonstrate that the complexes known to be involved in RNA synthesis contain multiple gene 1 proteins and are closely associated with structural proteins at presumed sites of virion assembly.