In eukaryotes, cap-dependent translation initiation is a sophisticated process that requires numerous trans-acting factors, the eukaryotic Initiation Factors (eIFs). Their main function is to assist the ribosome for accurate AUG start codon recognition. The whole process requires a 5'-3' scanning step and is therefore highly dynamic. Therefore translation requires a complex interplay between eIFs through assembly/release cycles. Here, we describe an original approach to assess the dynamic features of translation initiation. The principle is to use the m7Gcap located at the 5' extremity of mRNAs as a tracker to monitor RNA and protein components that are in its vicinity. Cap-binding molecules are trapped by chemical and UV crosslinking. The combination of cap crosslinking methods in cell-free translation systems with the use of specific translation inhibitors for different steps such as edeine, GMP-PNP or cycloheximide allowed assessing the cap fate during eukaryotic translation. Here, we followed the position of the cap in the histone H4 mRNA and the cap binding proteins during H4 mRNA translation.

中文翻译：

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通过交联方法在翻译起始过程中跟踪 m 7 G-cap

在真核生物中，帽依赖性翻译起始是一个复杂的过程，需要许多反式作用因子，即真核起始因子 (eIF)。它们的主要功能是协助核糖体准确识别 AUG 起始密码子。整个过程需要 5'-3' 扫描步骤，因此是高度动态的。因此，翻译需要通过组装/发布周期在 eIF 之间进行复杂的相互作用。在这里，我们描述了一种评估翻译起始动态特征的原始方法。其原理是使用位于 mRNA 5' 末端的 m7Gcap 作为跟踪器来监测其附近的 RNA 和蛋白质成分。帽结合分子被化学和紫外线交联捕获。无细胞翻译系统中的帽交联方法与不同步骤（如 edeine、GMP-PNP 或放线菌酮）使用特定翻译抑制剂的组合允许评估真核翻译过程中的帽命运。在这里，我们在 H4 mRNA 翻译过程中跟踪了组蛋白 H4 mRNA 中帽的位置和帽结合蛋白。