Herpes Simplex Virus Type 1 ICP0 Phosphorylation Mutants Impair the E3 Ubiquitin Ligase Activity of ICP0 in a Cell Type-Dependent Manner
Herpes Simplex Virus Type 1 ICP0 Phosphorylation Mutants Impair the E3 Ubiquitin Ligase Activity of ICP0 in a Cell Type-Dependent Manner
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DOI：
10.1128/jvi.01063-08
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发表时间：
2008-11-01
影响因子：
5.4
通讯作者：
Davido, David
中科院分区：
文献类型：
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作者：
Boutell, Chris;Everett, Roger;Davido, David
单纯疱疹病毒1型（HSV-1）感染细胞蛋白0（ICP 0）是一种110 kDa的核磷蛋白，是有效启动裂解感染和从潜伏期重新激活静止病毒基因组所必需的。ICP 0作为一种有效的病毒反式激活因子的能力是由其N端锌结合环指结构域介导的。该结构域赋予ICP 0 E3泛素连接酶活性，并且是感染期间许多细胞蛋白的蛋白酶体依赖性降解所需的，包括主要核结构域10（ND 10）组成蛋白早幼粒细胞白血病。在先前的工作中，我们绘制了ICP 0内的三个磷酸化区域，其中两个直接影响其在瞬时转染测定中的反式激活能力（Davido等人，J. Virol. 79：1232-1243，2005）。由于ICP 0是一种磷蛋白，我们最初试图测试磷酸化调节ICP 0的E3泛素连接酶活性的假设。虽然没有一个突变影响ICP 0 E3连接酶的体外活性，瞬时转染分析表明，一个或多个磷酸化区域内的突变损害了ICP 0与共定位缀合的泛素形成病灶的能力，并破坏ND 10。其中一个区域内的突变也影响ICP 0的稳定性，所有这些现象都以细胞类型依赖性方式发生。在病毒感染的背景下，与所有其他测试的病毒相比，只有一种ICP 0磷酸化突变体（P1）显示出病毒复制的显著缺陷和增强的蛋白质稳定性。这项研究表明，特定的细胞环境和表达的背景下（转染与感染）差异调节ICP 0相关的E3泛素连接酶活性通过磷酸化的几个活动。
Herpes simplex virus type 1 (HSV-1) infected cell protein 0 (ICP0) is a 110-kDa nuclear phosphoprotein that is required for both the efficient initiation of lytic infection and the reactivation of quiescent viral genomes from latency. The ability of ICP0 to act as a potent viral transactivator is mediated by its N-terminal zinc-binding RING finger domain. This domain confers E3 ubiquitin ligase activity to ICP0 and is required for the proteasome-dependent degradation of a number of cellular proteins during infection, including the major nuclear domain 10 (ND10) constituent protein promyelocytic leukemia. In previous work we mapped three phosphorylation regions within ICP0, two of which directly affected its transactivation capabilities in transient transfection assays (Davido et al., J. Virol. 79: 1232-1243, 2005). Because ICP0 is a phosphoprotein, we initially sought to test the hypothesis that phosphorylation regulates the E3 ubiquitin ligase activity of ICP0. Although none of the mutations affected ICP0 E3 ligase activity in vitro, transient transfection analysis indicated that mutations within one or more of the phosphorylated regions impaired the ability of ICP0 to form foci with colocalizing conjugated ubiquitin and to disrupt ND10. Mutations within one of the regions also affected ICP0 stability, and all of these phenomena occurred in a cell type-dependent manner. In the context of viral infection, only one ICP0 phosphorylation mutant (P1) showed a significant defect in viral replication and enhanced protein stability compared to all the other viruses tested. This study suggests that specific cellular environments and context of expression (transfection versus infection) differentially regulate several activities of ICP0 related to its E3 ubiquitin ligase activity via phosphorylation.