EFFECT OF SEQUENCE CONTEXT AT STOP CODONS ON EFFICIENCY OF REINITIATION IN GCN4 TRANSLATIONAL CONTROL
EFFECT OF SEQUENCE CONTEXT AT STOP CODONS ON EFFICIENCY OF REINITIATION IN GCN4 TRANSLATIONAL CONTROL
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DOI：
10.1128/mcb.14.1.606
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发表时间：
1994-01-01
影响因子：
5.3
通讯作者：
HINNEBUSCH, AG
中科院分区：
文献类型：
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作者：
GRANT, CM;HINNEBUSCH, AG
GCN 4基因的翻译控制涉及mRNA前导序列中的两个短的开放阅读框（uORF 1和uORF 4），它们在自身翻译后允许GCN 4重新起始的能力方面存在很大差异。uORF 4的低效率再起始特征可以在杂交元件中重建，其中uORF 1的最后一个密码子和其终止密码子3'端的10个核苷酸（终止区）被来自uORF 4的相应核苷酸取代。为了确定这13个核苷酸的特征，确定它们对再起始的影响，我们分别随机化了uORF 1-uORF 4杂合体的第三密码子和终止区的序列，并选择了具有uORF 1特征的高水平再起始的突变等位基因。结果表明，许多不同的A+ U丰富的三联体存在于uORF 1的第三个密码子可以克服来自uOPF 4的终止区对GCN 4的再起始效率的抑制作用。有效的再起始与指定特定氨基酸或异受体tRNA的密码子无关。类似地，我们发现，在uORF 1的终止区域中存在的A+ U丰富序列的多样性集合可以在来自uORF 4的第三个密码子的存在下恢复在GCN 4处的有效再起始。为了解释这些结果，我们建议，重新启动可以受损的稳定碱基配对之间的核苷酸侧翼的uORF 1终止密码子和tRNA的配对与第三个密码子，rRNA，或位于GCN 4 mRNA的其他地方的序列。我们认为，这些相互作用延迟恢复扫描后，在uORF的肽链终止，从而导致核糖体从mRNA的解离。
Translational control of the GCN4 gene involves two short open reading frames in the mRNA leader (uORF1 and uORF4) that differ greatly in the ability to allow reinitiation at GCN4 following their own translation. The low efficiency of reinitiation characteristic of uORF4 can be reconstituted in a hybrid element in which the last codon of uORF1 and 10 nucleotides 3' to its stop codon (the termination region) are substituted with the corresponding nucleotides from uORF4. To define the features of these 13 nucleotides that determine their effects on reinitiation, we separately randomized the sequence of the third codon and termination region of the uORF1-uORF4 hybrid and selected mutant alleles with the high-level reinitiation that is characteristic of uORF1. The results indicate that many different A+U-rich triplets present at the third codon of uORF1 can overcome the inhibitory effect of the termination region derived from uOPF4 on the efficiency of reinitiation at GCN4. Efficient reinitiation is not associated with codons specifying a particular amino acid or isoacceptor tRNA. Similarly, we found that a diverse collection of A+U-rich sequences present in the termination region of uORF1 could restore efficient reinitiation at GCN4 in the presence of the third codon derived from uORF4. To explain these results, we propose that reinitiation can be impaired by stable base pairing between nucleotides flanking the uORF1 stop codon and either the tRNA which pairs with the third codon, the rRNA, or sequences located elsewhere in GCN4 mRNA. We suggest that these interactions delay the resumption of scanning following peptide chain termination at the uORF and thereby lead to ribosome dissociation from the mRNA.