Plant Pathology Department, and Biochemistry, Biophysics, and Molecular Biology Department, Iowa State University, Ames, IA 50011, USA.
Structure. 2011 Jun 8;19(6):868-80. doi: 10.1016/j.str.2011.03.013.
Eukaryotic initiation factor eIF4E performs a key early step in translation by specifically recognizing the m⁷GpppN cap structure at the 5' end of cellular mRNAs. Many viral mRNAs lack a 5' cap and thus bypass eIF4E. In contrast, we reported a cap-independent translation element (PTE) in Pea enation mosaic virus RNA2 that binds and requires eIF4E for translation initiation. To understand how this uncapped RNA is bound tightly by eIF4E, we employ SHAPE probing, phylogenetic comparisons with new PTEs discovered in panico- and carmoviruses, footprinting of the eIF4E binding site, and 3D RNA modeling using NAST, MC-Fold, and MC-Sym to predict a compact, 3D structure of the RNA. We propose that the cap-binding pocket of eIF4E clamps around a pseudoknot, placing a highly SHAPE-reactive guanosine in the pocket in place of the normal m⁷GpppN cap. This reveals a new mechanism of mRNA recognition by eIF4E.
真核起始因子 eIF4E 通过特异性识别细胞 mRNA 5'端的 m⁷GpppN 帽结构，在翻译中发挥关键的早期步骤。许多病毒 mRNA 缺乏 5'帽结构，因此绕过了 eIF4E。相比之下，我们报道了 Pea enation mosaic virus RNA2 中的一个无帽依赖翻译元件（PTE），它结合并需要 eIF4E 来启动翻译。为了了解这种无帽 RNA 如何被 eIF4E 紧密结合，我们采用了 SHAPE 探测、与在 Panico 和 Carmoviruses 中发现的新 PTEs 的系统发育比较、eIF4E 结合位点的足迹分析，以及使用 NAST、MC-Fold 和 MC-Sym 进行的 3D RNA 建模，以预测 RNA 的紧凑 3D 结构。我们提出，eIF4E 的帽结合口袋夹住假结，将高度 SHAPE 反应性的鸟苷置于口袋中，取代正常的 m⁷GpppN 帽。这揭示了 eIF4E 识别 mRNA 的一种新机制。
