Step-wise formation of eukaryotic double-row polyribosomes and circular translation of polysomal mRNA
Step-wise formation of eukaryotic double-row polyribosomes and circular translation of polysomal mRNA
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DOI：
10.1093/nar/gkm1177
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发表时间：
2008-06-01
影响因子：
14.9
通讯作者：
Spirin, Alexander S.
中科院分区：
文献类型：
--
作者：
Kopeina, Gelina S.;Afonina, Zhanna A.;Spirin, Alexander S.
使用沉降和电子显微镜技术在长期小麦胚芽无细胞翻译系统中研究了多核糖体形成的时间过程。多聚核糖体在未加帽的荧光素酶 mRNA 上形成，具有翻译增强的 5 和 3 UTR。发现满载多核糖体的形成是一个漫长的过程，需要多轮翻译并经历几个阶段。首先，形成含有不超过六个核糖体的短线性多聚核糖体。接下来，这些多核糖体折叠成短的双行簇。随后簇的逐渐伸长产生了含有多达 3040 个核糖体的重负载双排串。双排多核糖体的形成被认为相当于多核糖体的环化，其中反平行的半个环通过核糖体相互作用横向粘在一起。在双排型多核糖体中观察到的与游离核糖体和游离mRNA的缓慢交换，以及它们对AMP-PNP的翻译抗性，提供了证据表明大多数多核糖体在环化多核糖体内重新启动翻译而无需扫描5 UTR，而包括5 UTR扫描的从头启动以慢得多的速度进行。 5和3 UTR的去除或替换影响了翻译的初始阶段，但并没有阻止翻译过程中双排多核糖体的形成。
The time course of polysome formation was studied in a long-term wheat germ cell-free translation system using sedimentation and electron microscopy techniques. The polysomes were formed on uncapped luciferase mRNA with translation-enhancing 5 and 3 UTRs. The formation of fully loaded polysomes was found to be a long process that required many rounds of translation and proceeded via several phases. First, short linear polysomes containing no more than six ribosomes were formed. Next, folding of these polysomes into short double-row clusters occurred. Subsequent gradual elongation of the clusters gave rise to heavy-loaded double-row strings containing up to 3040 ribosomes. The formation of the double-row polysomes was considered to be equivalent to circularization of polysomes, with antiparallel halves of the circle being laterally stuck together by ribosome interactions. A slow exchange with free ribosomes and free mRNA observed in the double-row type polysomes, as well as the resistance of translation in them to AMP-PNP, provided evidence that most polysomal ribosomes reinitiate translation within the circularized polysomes without scanning of 5 UTR, while de novo initiation including 5 UTR scanning proceeds at a much slower rate. Removal or replacements of 5 and 3 UTRs affected the initial phase of translation, but did not prevent the formation of the double-row polysomes during translation.