Structure and function of a cap-independent translation element that functions in either the 3′ or the 5′ untranslated region
Structure and function of a cap-independent translation element that functions in either the 3′ or the 5′ untranslated region
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DOI：
10.1017/s1355838200001539
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发表时间：
2000-12-01
期刊：
影响因子：
4.5
通讯作者：
Miller, WA
中科院分区：
文献类型：
--
作者：
Guo, L;Allen, E;Miller, WA
大麦黄矮病毒RNA缺乏5'帽和poly(A)尾，但它的翻译效率很高。它包含位于 3' UTR 中的帽独立翻译元件 (TE)，可在最接近 mRNA 5' 末端的 AUG 处提供有效的翻译起始。我们建议 TE 必须既招募核糖体又促进 3'-5' 通讯。为了剖析其功能，我们确定了 TE 的二级结构及其中域的作用。核酸酶探测和结构定向诱变表明，105-nt TE (TE105) 形成十字形二级结构，其中包含由单链区域连接的四个螺旋。 TE105 可以在小麦胚芽翻译提取物的任一 UTR 中发挥作用。植物细胞中完全不依赖帽子的翻译需要更长的病毒序列（最多 869 nt）。然而，使用 TE105 与 Poly(A) 尾结合，可以在植物细胞中获得未加帽的 mRNA 的大量翻译。所有二级结构元件和大多数突变的一级序列都是 3' 和 5' UTR 环境中帽独立翻译所必需的。仅在 3' UTR 上下文中才需要七碱基环序列。因此，该环序列可能仅参与 UTR 之间的通讯，而不直接参与招募翻译机制。这种结构和功能分析提供了一个框架，用于理解一类新兴的与帽无关的翻译元素，其特征在于它们在 3' UTR 中的位置。
Barley yellow dwarf virus RNA lacks both a 5' cap and a poly(A) tail, yet it is translated efficiently. It contains a cap-independent translation element (TE), located in the 3' UTR, that confers efficient translation initiation at the AUG closest to the 5' end of the mRNA. We propose that the TE must both recruit ribosomes and facilitate 3'-5' communication. To dissect its function, we determined the secondary structure of the TE and roles of domains within it. Nuclease probing and structure-directed mutagenesis revealed that the 105-nt TE (TE105) forms a cruciform secondary structure containing four helices connected by single-stranded regions. TE105 can function in either UTR in wheat germ translation extracts. A longer viral sequence (at most 869 nt) is required for full cap-independent translation in plant cells. However, substantial translation of uncapped mRNAs can be obtained in plant cells with TE105 combined with a poly(A) tail. All secondary structural elements and most primary sequences that were mutated are required for cap-independent translation in the 3' and 5' UTR contexts. A seven-base loop sequence was needed only in the 3' UTR context. Thus, this loop sequence may be involved only in communication between the UTRs and not directly in recruiting translational machinery. This structural and functional analysis provides a framework for understanding an emerging class of cap-independent translation elements distinguished by their location in the 3' UTR.