The position dependence of translational regulation via RNA-RNA and RNA-protein interactions in the 5'-untranslated region of eukaryotic mRNA is a function of the thermodynamic competence of 40 S ribosomes in translational initiation
The position dependence of translational regulation via RNA-RNA and RNA-protein interactions in the 5'-untranslated region of eukaryotic mRNA is a function of the thermodynamic competence of 40 S ribosomes in translational initiation
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DOI：
10.1074/jbc.272.26.16531
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发表时间：
1997-06-27
影响因子：
4.8
通讯作者：
McCarthy, JEG
中科院分区：
文献类型：
--
作者：
Koloteva, N;Muller, PP;McCarthy, JEG
为了使二级结构和rna结合蛋白能够通过哺乳动物mrna的5'-非翻译区（5'-UTR）抑制翻译起始，Cap邻近是一个必要条件。我们发现，在酿酒酵母中，与哺乳动物细胞不同，哺乳动物铁调控蛋白1 （IRP1）的体外翻译抑制作用与它在5'-UTR中的靶位点铁响应元件（IRE）无关。体外研究表明，IRP1的结合亲和力也不受IRE位置的影响。使用IRE环突变体，我们观察到酵母中irp1依赖性抑制几乎完全丧失，同时对IRE靶点的结合亲和力降低了150倍。这反映了铁诱导的哺乳动物细胞中IRE/IRP1亲和力调节的自然定量范围。通过增强IRE茎环的稳定性，我们还发现其固有的折叠能与IRP1的结合能一起作用，从而提供了限制翻译起始的附加能力。酵母中位于帽端末端位置的IRE-IRP1复合物阻断了对5'-UTR上40 S核糖体的扫描。由此可见，哺乳动物位点特异性翻译抑制的位置效应是由哺乳动物起始前复合物在起始过程的不同步骤中破坏抑制结构的能力决定的。
Cap proximity is a requirement to enable secondary structures and RNA-binding proteins to repress translational initiation via the 5'-untranslated region (5'-UTR) of mammalian mRNAs. We show that in Saccharomyces cerevisiae, unlike mammalian cells, the in vitro translational repressive effect of the mammalian iron regulatory protein 1 (IRP1) is independent of the site of its target in the 5'-UTR, the iron-responsive element (IRE). In vitro studies demonstrate that the binding affinity of IRP1 is also unaffected by the position of the IRE. Using IRE loop mutants, we observe an almost complete loss of IRP1-dependent repression in yeast concomitant with a 150-fold reduction in binding affinity for the IRE target. This mirrors the natural quantitative range of iron-induced adjustment of IRE/IRP1 affinity in mammalian cells. By enhancing the stability of the IRE stem-loop, we also show that its intrinsic folding energy acts together with the binding energy of IRP1 to give an additive capacity to restrict translational initiation. An IRE-IRP1 complex in a cap-distal position in yeast blocks scanning 40 S ribosomes on the 5'-UTR, It follows that the position effect of mammalian site-specific translational repression is dictated by the competence of the mammalian preinitiation complex to destabilize inhibitory structures at different steps of the initiation process.