RNA. 2014 Jan;20(1):1-8. doi: 10.1261/rna.038182.113. Epub 2013 Nov 19.
The HIV-1 Rev protein mediates export of unspliced and singly spliced viral transcripts by binding to the Rev response element (RRE) and recruiting the cellular export factor CRM1. Here, we investigated the recruitment of Rev to the transcription sites of HIV-1 reporters that splice either post- or cotranscriptionally. In both cases, we observed that Rev localized to the transcription sites of the reporters and recruited CRM1. Rev and CRM1 remained at the reporter transcription sites when cells were treated with the splicing inhibitor Spliceostatin A (SSA), showing that the proteins associate with RNA prior to or during early spliceosome assembly. Fluorescence recovery after photobleaching (FRAP) revealed that Rev and CRM1 have similar kinetics as the HIV-1 RNA, indicating that Rev, CRM1, and RRE-containing RNAs are released from the site of transcription in one single export complex. These results suggest that cotranscriptional formation of a stable export complex serves as a means to ensure efficient export of unspliced viral RNAs.
HIV-1 Rev 蛋白通过结合 Rev 反应元件（RRE）并招募细胞输出因子 CRM1，介导未剪接和单剪接病毒转录本的输出。在这里，我们研究了 Rev 对 HIV-1 报告基因转录本的募集情况，这些报告基因要么在转录后剪接，要么在共转录时剪接。在这两种情况下，我们都观察到 Rev 定位于报告基因转录本的位置，并招募 CRM1。当用剪接抑制剂 Spliceostatin A（SSA）处理细胞时，Rev 和 CRM1 仍留在报告基因转录本的位置，这表明这些蛋白质在剪接体组装之前或早期与 RNA 结合。光漂白后荧光恢复（FRAP）显示 Rev 和 CRM1 的动力学与 HIV-1 RNA 相似，这表明 Rev、CRM1 和含有 RRE 的 RNA 从转录位点以一个单一的输出复合物释放。这些结果表明，共转录形成稳定的输出复合物是确保未剪接病毒 RNA 有效输出的一种手段。
