Unbiased selection of localization elements reveals cis-acting determinants of mRNA bud localization in Saccharomyces cerevisiae
Unbiased selection of localization elements reveals cis-acting determinants of mRNA bud localization in Saccharomyces cerevisiae
复制标题
DOI：
10.1073/pnas.0509229102
复制
发表时间：
2005-12-13
影响因子：
11.1
通讯作者：
DeRisi, JL
中科院分区：
文献类型：
--
作者：
Jambhekar, A;McDermott, K;DeRisi, JL
细胞质 mRNA 定位是许多生物体用来产生不对称性和隔离蛋白质活性的机制。在酿酒酵母中，mRNA 向分裂细胞芽尖的转运是通过 She2p、She3p 和 Myo4p 与 RNA 编码区的结合介导的。迄今为止，已鉴定出 24 个芽定位 mRNA，但介导定位的 RNA 决定簇仍知之甚少。在这里，我们使用非同源随机重组来生成序列文库，可以根据其结合 She 复合物蛋白的能力进行选择，从而提供一种公正的方法来最小化和绘制几种转运 RNA 中的定位元件。对衍生序列和预测的二级结构的分析揭示了介导与 She 复合物的结合以及体内 RNA 定位于芽尖的短序列基序。预测的单链核心 CG 二核苷酸似乎是 RNA-蛋白质界面的重要组成部分，尽管其他核苷酸以上下文相关的方式发挥作用。我们的发现进一步加深了我们对 She 复合物的 RNA 识别的理解，这里使用的方法应该适用于阐明参与许多其他类型相互作用的最小 RNA 基序。
Cytoplasmic mRNA localization is a mechanism used by many organisms to generate asymmetry and sequester protein activity. In the yeast Saccharomyces cerevisiae, mRNA transport to bud tips of dividing cells is mediated by the binding of She2p, She3p, and Myo4p to coding regions of the RNA. To date, 24 bud-localized mRNAs have been identified, yet the RNA determinants that mediate localization remain poorly understood. Here, we used nonhomologous random recombination to generate libraries of sequences that could be selected for their ability to bind She-complex proteins, thereby providing an unbiased approach for minimizing and mapping localization elements in several transported RNAs. Analysis of the derived sequences and predicted secondary structures revealed short sequence motifs that mediate binding to the She complex and RNA localization to the bud tip in vivo. A predicted single-stranded core CG dinucleotide appears to be an important component of the RNA-protein interface, although other nucleotides contribute in a context-dependent manner. Our findings further our understanding of RNA recognition by the She complex, and the methods used here should be applicable for elucidating minimal RNA motifs involved in many other types of interactions.