Jain C, Belasco J G
Skirball Institute of Biomolecular Medicine and Department of Microbiology, New York University School of Medicine, New York 10016, USA.
Mol Cell. 2001 Mar;7(3):603-14. doi: 10.1016/s1097-2765(01)00207-6.
The functional efficacy of the HIV-1 Rev protein is highly dependent on its ability to assemble onto its HIV-1 RNA target (the RRE) as a multimeric complex. To elucidate the mechanism of multimeric assembly, we have devised two rapid and broadly applicable strategies for examining cooperative interactions between proteins bound to RNA, one based on cooperative translational repression of a two-site reporter and the other on gel shift analysis with crude E. coli extracts. Using these strategies, we have identified two distinct surfaces of Rev (head and tail) that are critical for different steps in multimeric assembly. Our data indicate that Rev assembles cooperatively on the RRE via a series of symmetrical tail-to-tail and head-to-head protein-protein interactions. The insights into molecular architecture suggested by these findings have enabled us to derive a structural model for Rev and its multimerization on the RRE.
HIV-1 Rev蛋白的功能效力高度依赖于其以多聚体复合物形式组装到HIV-1 RNA靶标（RRE）上的能力。为阐明多聚体组装机制，我们设计了两种快速且广泛适用的策略来检测与RNA结合的蛋白质之间的协同相互作用，一种基于对双位点报告基因的协同翻译抑制，另一种基于使用大肠杆菌粗提物的凝胶迁移分析。利用这些策略，我们确定了Rev的两个不同表面（头部和尾部），它们对于多聚体组装的不同步骤至关重要。我们的数据表明，Rev通过一系列对称的尾对尾和头对头蛋白质-蛋白质相互作用在RRE上协同组装。这些发现所揭示的分子结构见解使我们能够推导出Rev及其在RRE上多聚化的结构模型。
