Peng Rongsheng, Moses Stephanie C, Tan Jie, Kremmer Elisabeth, Ling Paul D
Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, TX 77030, USA.
J Virol. 2005 Apr;79(7):4492-505. doi: 10.1128/JVI.79.7.4492-4505.2005.
The mechanistic contribution of the Epstein-Barr virus (EBV) EBNA-LP protein to B-cell immortalization remains an enigma. However, previous studies have indicated that EBNA-LP may contribute to immortalization by enhancing EBNA2-mediated transcriptional activation of the LMP-1 gene. To gain further insight into the potential role EBNA-LP has in EBV-mediated B-cell immortalization, we asked whether it is a global or gene-specific coactivator of EBNA2 and whether coactivation requires interaction between these proteins. In type I Burkitt's lymphoma cells, we found that EBNA-LP strongly coactivated EBNA2 stimulation of LMP-1 and LMP2B RNAs, which are expressed from the viral divergent promoter. Surprisingly, the viral LMP2A gene and cellular CD21 and Hes-1 genes were induced by EBNA2 but showed no further induction after EBNA-LP coexpression. We also found that EBNA-LP did not stably interact with EBNA2 in coimmunoprecipitation assays, even though the conditions were adequate to observe specific interactions between EBNA2 and its cellular cofactor, CBF1. Colocalization between EBNA2 and EBNA-LP was not detectable in EBV-transformed cell lines or transfected type I Burkitt's cells. Finally, no significant interactions between EBNA2 and EBNA-LP were found with mammalian two-hybrid assays. From this data, we conclude that EBNA-LP is not a global coactivator of EBNA2 targets, but it preferentially coactivates EBNA2 stimulation of the viral divergent promoter. While this may require specific transient interactions between these proteins that only occur in the context of the divergent promoter, our data strongly suggest that EBNA-LP also cooperates with EBNA2 through mechanisms that do not require direct or indirect complex formation between these proteins.
爱泼斯坦-巴尔病毒（EBV）的EBNA-LP蛋白对B细胞永生化的机制性贡献仍是一个谜。然而，先前的研究表明，EBNA-LP可能通过增强EBNA2介导的LMP-1基因转录激活来促进永生化。为了进一步深入了解EBNA-LP在EBV介导的B细胞永生化中的潜在作用，我们研究了它是EBNA2的全局性还是基因特异性共激活因子，以及共激活是否需要这些蛋白之间的相互作用。在I型伯基特淋巴瘤细胞中，我们发现EBNA-LP强烈共激活EBNA2对LMP-1和LMP2B RNA的刺激，这两种RNA由病毒差异启动子表达。令人惊讶的是，病毒LMP2A基因以及细胞CD21和Hes-1基因由EBNA2诱导，但在共表达EBNA-LP后没有进一步诱导。我们还发现，在免疫共沉淀试验中，EBNA-LP与EBNA2没有稳定的相互作用，尽管条件足以观察到EBNA2与其细胞辅因子CBF1之间的特异性相互作用。在EBV转化的细胞系或转染的I型伯基特细胞中未检测到EBNA2和EBNA-LP之间的共定位。最后，在哺乳动物双杂交试验中未发现EBNA2和EBNA-LP之间有显著相互作用。根据这些数据，我们得出结论，EBNA-LP不是EBNA2靶标的全局性共激活因子，但它优先共激活EBNA2对病毒差异启动子的刺激。虽然这可能需要这些蛋白之间仅在差异启动子背景下发生的特异性瞬时相互作用，但我们的数据强烈表明，EBNA-LP也通过不需要这些蛋白之间直接或间接形成复合物的机制与EBNA2协同作用。
