Herpes Simplex Virus 1 Protein Kinase Us3 and Major Tegument Protein UL47 Reciprocally Regulate Their Subcellular Localization in Infected Cells
Herpes Simplex Virus 1 Protein Kinase Us3 and Major Tegument Protein UL47 Reciprocally Regulate Their Subcellular Localization in Infected Cells
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DOI：
10.1128/jvi.00845-11
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发表时间：
2011-09-01
影响因子：
5.4
通讯作者：
Kawaguchi, Yasushi
中科院分区：
文献类型：
--
作者：
Kato, Akihisa;Liu, Zhuoming;Kawaguchi, Yasushi
Us 3是由单纯疱疹病毒1（HSV-1）编码的丝氨酸-苏氨酸蛋白激酶。我们已经确定了UL 47，一个主要的病毒体蛋白，作为一种新的生理底物的Us 3。在体外激酶测定和突变的系统分析，在推定的Us 3磷酸化位点附近的核定位信号的UL 47显示，丝氨酸残基77（Ser-77）的Us 3磷酸化的UL 47所需的。用丙氨酸替换UL 47 Ser-77导致UL 47在核边缘的异常积累，并损害了UL 47在感染细胞的显著部分中的核定位。UL 47定位中的相同缺陷是由Us 3中的氨基酸取代产生的，该氨基酸取代使其蛋白激酶活性失活。相比之下，在UL 47 Ser-77磷酸化突变恢复野生型核定位。UL 47 S77 A突变还减少了小鼠角膜中的病毒复制和小鼠疱疹性角膜基质炎的发展。此外，UL 47在感染细胞中与Us 3形成稳定的复合物，并且在不存在UL 47的情况下Us 3的核定位显著受损。这些结果表明，UL 47 Ser-77的Us 3磷酸化促进了UL 47在细胞培养物中的核定位，并且在体内病毒复制和致病中起关键作用。此外，UL 47似乎是感染细胞中Us 3的有效核定位所需的。因此，Us 3蛋白激酶及其底物UL 47表现出独特的调节特征，因为它们在感染细胞中可调节其亚细胞定位。
Us3 is a serine-threonine protein kinase encoded by herpes simplex virus 1 (HSV-1). We have identified UL47, a major virion protein, as a novel physiological substrate of Us3. In vitro kinase assays and systematic analysis of mutations at putative Us3 phosphorylation sites near the nuclear localization signal of UL47 showed that serine at residue 77 (Ser-77) was required for Us3 phosphorylation of UL47. Replacement of UL47 Ser-77 by alanine produced aberrant accumulation of UL47 at the nuclear rim and impaired the nuclear localization of UL47 in a significant fraction of infected cells. The same defect in UL47 localization was produced by an amino acid substitution in Us3 that inactivated its protein kinase activity. In contrast, a phosphomimetic mutation at UL47 Ser-77 restored wild-type nuclear localization. The UL47 S77A mutation also reduced viral replication in the mouse cornea and the development of herpes stromal keratitis in mice. In addition, UL47 formed a stable complex with Us3 in infected cells, and nuclear localization of Us3 was significantly impaired in the absence of UL47. These results suggested that Us3 phosphorylation of UL47 Ser-77 promoted the nuclear localization of UL47 in cell cultures and played a critical role in viral replication and pathogenesis in vivo. Furthermore, UL47 appeared to be required for efficient nuclear localization of Us3 in infected cells. Therefore, Us3 protein kinase and its substrate UL47 demonstrated a unique regulatory feature in that they reciprocally regulated their subcellular localization in infected cells.