The gene 10 (UL49.5) product of equine herpesvirus 1 is necessary and sufficient for functional processing of glycoprotein M
The gene 10 (UL49.5) product of equine herpesvirus 1 is necessary and sufficient for functional processing of glycoprotein M
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DOI：
10.1128/jvi.76.6.2952-2963.2002
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发表时间：
2002-03-01
影响因子：
5.4
通讯作者：
Osterrieder, N
中科院分区：
文献类型：
--
作者：
Rudolph, J;Seyboldt, C;Osterrieder, N
分析了马疱疹病毒I型（EHV-1）糖蛋白NI（gM）与基因10（UL49.5）产物的功能协同作用。使用gM和UL49.5表达质粒以及组成型表达UL49.5（RK49.5）或gM（RKgM）的RK 13细胞系的瞬时转染实验证明，仅在共表达两种蛋白质后才可检测到gM的内-β-N-乙酰氨基葡糖苷酶H（endo If）抗性成熟形式。在KyA株中EHV-1 UL49.5同源基因10的缺失导致小空斑表型和高达190倍的病毒滴度降低。突变体KyADelta49.5病毒的生长缺陷与GM阴性KyA病毒的生长缺陷非常相似，可以通过突变体病毒在RK49.5细胞上的生长或通过修复回复突变体病毒KyADelta49.5R中基因10的缺失来完全补偿。用UL49.5阴性EHV-1感染的细胞的分析表明，在不存在UL49.5产物的情况下，gM不被转运到trans-Golgi网络。相比之下，在KyADelta49.5感染的RK49.5细胞中，gM被有效转运并加工成内H抗性成熟形式。此外，放射免疫沉淀实验表明，只有当10，000-M-r蛋白与KyA或KyADelta 49. 5 R感染的细胞或病毒体中的gM共沉淀时，才观察到gM成熟。该蛋白在感染KyDelta49.5或KyADeltagM的细胞中不存在，表明它是EHV-1 UL49.5产物。综上所述，我们的结果表明，EHV-1 UL49.5产物的表达对于gM加工是必要的和足够的，并且是有效病毒复制所需的。
The functional cooperation of equine herpesvirus I (EHV-1) glycoprotein NI (gM) and the gene 10 (UL49.5) product was analyzed. Transient-transfection experiments using gM and UL49.5 expression plasmids as well as RK13 cell lines constitutively expressing UL49.5 (RK49.5) or gM (RKgM) demonstrated that the endo-beta-N-acetylglucosaminidase H (endo If)-resistant mature form of gM was detectable only after coexpression of the two proteins. Deletion of the EHV-1 UL49.5-homologous gene 10 in strain KyA resulted in a small-plaque phenotype and up to 190-fold-reduced virus titers. The growth defects of the mutant KyADelta49.5 virus, which were very similar to those of a gM-negative KyA virus, could be completely compensated for by growth of the mutant virus on RK49.5 cells or by repairing the deletion of gene 10 in the revertant virus KyADelta49.5R. Analysis of cells infected with the UL49.5-negative EHV-1 demonstrated that gM was not transported to the trans-Golgi network in the absence of the UL49.5 product. In contrast, gM was efficiently transported and processed to the endo H-resistant mature form in KyADelta49.5-infected RK49.5 cells. Furthermore, radioimmunoprecipitation experiments demonstrated that gM maturation was observed only if a 10,000-M-r protein was coprecipitated with gM in KyA- or KyADelta49.5R-infected cells or virions. This protein was absent in cells infected with KyDelta49.5 or KyADeltagM, suggesting that it was the EHV-1 UL49.5 product. Taken together, our results demonstrate that the expression of the EHV-1 UL49.5 product is necessary and sufficient for gM processing and that it is required for efficient virus replication.