Papugani Anil, Coleman Tricia, Jones Clinton, Zhang Luwen
School of Biological Sciences, University of Nebraska, Lincoln, NE 68588, USA.
Virus Res. 2008 Jan;131(1):1-7. doi: 10.1016/j.virusres.2007.07.019. Epub 2007 Sep 11.
Kaposi's sarcoma-associated herpesvirus (KSHV) replication and transcription activator (RTA) is necessary and sufficient for the switch from KSHV latency to lytic replication. RTA activates promoters by several mechanisms. RTA can bind to sequences in viral promoters and activate transcription. In addition, RTA interacts with the cellular recombination signal sequence-binding protein-J kappa (RBP-Jkappa), a transcriptional repressor, converts the repressor into an activator and activates viral promoters via RBP-Jkappa. Because RBP-Jkappa is required for RTA to activate lytic replication, it is important to understand how RTA cooperates with RBP-Jkappa protein to activate KSHV lytic replication program. Previously, we identified an RTA mutant, RTA-K152E, which has a defect in its direct DNA-binding activity. In this report, the effect of the mutant RTA on KSHV activation via RBP-Jkappa protein is examined. We demonstrate that RTA-K152E interacts with RBP-Jkappa physically and the mutant RTA and RBP-Jkappa complex binds to target DNA properly in vivo and in vitro. However, the complex of RTA-K152E and RBP-Jkappa does not activate transcription. Furthermore, the RTA mutant (RTA-K12E) inhibits cellular Notch-mediated RBP-Jkappa activation. These data collectively suggest that the complex between KSHV RTA and cellular RBP-Jkappa and the subsequent DNA binding by the complex are not sufficient for the activation of RBP-Jkappa protein. Other factor(s) whether additional cofactor(s) in the complex or the intrinsic conformation of RTA, are predicted to be required for the activation of RBP-Jkappa protein by RTA.
卡波西肉瘤相关疱疹病毒（KSHV）的复制和转录激活因子（RTA）对于KSHV从潜伏状态转换为裂解性复制是必需且充分的。RTA通过多种机制激活启动子。RTA可与病毒启动子中的序列结合并激活转录。此外，RTA与细胞重组信号序列结合蛋白-Jκ（RBP-Jκ）相互作用，RBP-Jκ是一种转录抑制因子，RTA将其转化为激活因子，并通过RBP-Jκ激活病毒启动子。由于RTA激活裂解性复制需要RBP-Jκ，因此了解RTA如何与RBP-Jκ蛋白协同激活KSHV裂解性复制程序很重要。此前，我们鉴定出一种RTA突变体RTA-K152E，其直接DNA结合活性存在缺陷。在本报告中，研究了突变型RTA通过RBP-Jκ蛋白对KSHV激活的影响。我们证明RTA-K152E与RBP-Jκ存在物理相互作用，并且突变型RTA与RBP-Jκ复合物在体内和体外均能正确结合靶DNA。然而，RTA-K152E与RBP-Jκ的复合物不能激活转录。此外，RTA突变体（RTA-K12E）抑制细胞Notch介导的RBP-Jκ激活。这些数据共同表明，KSHV RTA与细胞RBP-Jκ之间的复合物以及随后该复合物与DNA的结合不足以激活RBP-Jκ蛋白。预计RTA激活RBP-Jκ蛋白还需要其他因素，无论是复合物中的其他辅助因子还是RTA的内在构象。
