Post-translational modification of Rta of Epstein-Barr virus by SUMO-1
Post-translational modification of Rta of Epstein-Barr virus by SUMO-1
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DOI：
10.1074/jbc.m405470200
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发表时间：
2004-09-10
影响因子：
4.8
通讯作者：
Liu, ST
中科院分区：
文献类型：
--
作者：
Chang, LK;Lee, YH;Liu, ST
Epstein-Barr病毒(EBV)表达一种即刻早期蛋白RTA，激活EBV裂解基因的转录和裂解周期。本工作在酵母双杂交筛选中鉴定了Ubc9和PIAS1为RTA的结合伙伴。谷胱甘肽S转移酶下拉试验、免疫共沉淀和共聚焦显微镜证实了这些结合。这种相互作用似乎导致了RTA的总甲基化，因为不仅在体外RTA可以被总甲基化，而且在裂解诱导后的P3HR1细胞和在表达RTA和SUMO-1的质粒后的293T细胞中都可以检测到总甲基化的RTA。此外，PIAS1刺激SUMO-1与RTA的结合，从而起到E3连接酶的作用。此外，转染Ubc9、PIAS1和SUMO-1的表达载体可以增加RTA反式激活包含RTA反应元件的启动子的能力，这表明SUMO-1的修饰增加了RTA的反式激活活性。这项研究揭示了RTA在Lys-19、Lys-213和Lys-517残基上被求和，并且在Lys-19残基上的SUMO-1结合对于提高RTA的反式激活活性至关重要。这些结果表明，RTA的苏莫化可能在EBV裂解激活中起重要作用。
Epstein-Barr virus (EBV) expresses an immediate-early protein, Rta, to activate the transcription of EBV lytic genes and the lytic cycle. This work identifies Ubc9 and PIAS1 as binding partners of Rta in a yeast two-hybrid screen. These bindings are verified by glutathione S-transferase pull-down assay, coimmunoprecipitation, and confocal microscopy. The interactions appear to cause Rta sumoylation, because not only can Rta be sumoylated in vitro but also sumoylated Rta can be detected in P3HR1 cells following lytic induction and in 293T cells after transfecting plasmids that express Rta and SUMO-1. Moreover, PIAS1 stimulates conjugation of SUMO-1 to Rta, thus acting as an E3 ligase. Furthermore, transfecting plasmids that express Ubc9, PIAS1, and SUMO-1 increases the capacity of Rta to transactivate the promoter that includes an Rta response element, indicating that the modification by SUMO-1 increases the transactivation activity of Rta. This study reveals that Rta is sumoylated at the Lys-19, Lys-213, and Lys-517 residues and that SUMO-1 conjugation at the Lys-19 residue is crucial for enhancing the transactivation activity of Rta. These results indicate that sumoylation of Rta may be important in EBV lytic activation.